Increased expression of GAD mRNA during the chronic epileptic syndrome due to intrahippocampal tetanus toxin. A few mouse minimum lethal doses (MLD) of tetanus toxin injected into rat hippocampus triggers prolonged changes in neuronal function. Spontaneously recurring epileptic discharges arise in both the injected and the contralateral, uninjected hippocampus. The seizures remit after about 6 weeks, to be succeeded by a permanent depression of hippocampal neuronal responses. There is no evidence of any loss of pyramidal cells at this low dose of toxin. Here we studied presumptive inhibitory, GABAergic neurons, using in situ hybridization (ISH) with a probe directed against the mRNA encoding glutamic acid decarboxylase (GAD), at each of 1, 2, 4 and 8 weeks after injection of tetanus toxin. Epileptic activity was recorded from hippocampal slices prepared from both injected and contralateral hippocampi of rats at each time point, unexpectedly persisting until 8 weeks. There were no significant differences in the numbers of neurons containing GAD mRNA between toxin- and vehicle-injected and control rats in any hippocampal subfield, at any survival time, except for an apparently transient loss of hilar signal in vehicle-injected rats at 1 and 2 weeks which we attribute to a significant, transient loss of neuronal GAD mRNA to below the threshold for detection by ISH using this probe. In contrast there was a marked increase in GAD mRNA in the toxin-injected group, which reached a peak at 4 weeks, and returned to control levels by 8 weeks. The changes were bilateral and were most marked in the hilus of the dentate area, but were also significant in CA3 and CA1. Upregulation of GAD mRNA was preceded by an increase in the levels of the mRNA for the alpha subunit of the GTP binding protein, Gs (Gs alpha), at 2 weeks which affected the GABAergic neurons selectively, and not the pyramidal or granule cells. These marked changes in GAD mRNA may contribute to putative adaptive responses within GABAergic neurons, which would help contain epileptic activity in these chronic foci. The changes in GAD expression may be due to mechanisms acting through an increase in mRNA encoding Gs alpha.


Constancy of motor axon conduction time during growth in rats. Axon conduction distance, conduction velocity, and conduction time were measured for individual triceps surae motoneurons in Sprague-Dawley rats weighing 230-630 g (i.e., age range 6-16 weeks). Both conduction distance (nerve length) and velocity were closely correlated with weight (r = 0.95 and r = 0.82, respectively). In contrast, conduction time did not change as weight increased nearly threefold. This striking constancy is probably due to a corresponding increase in axon diameter. It could contribute to maintenance of stable motor performance during rapid growth.


Development of parvalbumin immunoreactive neurons in normal and intracranially transplanted retinas in the rat. Retinas from embryonic day 14 Sprague-Dawley rats were transplanted to the midbrain or cerebral cortex of newborn (P0) rats of which the right eye was enucleated at the time of transplantation. Parvalbumin immunoreactive (PV-I) neurons were studied in the developing retinal transplants, and in the remaining retina of the host, as well as in normal retinas. PV-I neurons were identifiable in retinas of normal and host rats from postnatal day 5 (P5) onward, with the PV-I somata primarily in the inner half of the inner nuclear layer and in the ganglion cell layer. An adult-like distribution of PV-I neurons was attained at P35, as judged by cell packing density, intensity of immunostaining, laminar distribution and soma size of subpopulations of PV-I cells. A similar time course of development and distribution of PV-I somata was observed in the retinal transplants, except for some minor differences such as a slight delay in PV-I cells achieving their final distribution. These findings provide evidence that PV-I neurons can survive, differentiate and mature according to predetermined programmes intrinsic to the retinal tissue following transplantation to a new and foreign environment.


Sprouting of fusimotor neurones after partial denervation of the cat soleus muscle. Normally, gamma motoneurones innervate only the intrafusal fibres of muscle spindles. This is a report of sprouting of gamma motoneurones to innervate extrafusal muscle fibres following partial denervation of the soleus muscle of kittens. In eight newborn animals, the L7 ventral root was cut on one side under anaesthesia and the animals were then allowed to recover. At approximately 100 days of age animals were reanaesthetised and a study made of mechanical properties of motor units whose axons ran in the S1 ventral root and supplied the partially denervated soleus muscle. Evidence was obtained for sprouting of all surviving alpha motoneurones. In addition, in four experiments axons conducting within the gamma range, on stimulation, produced measurable tension. In one experiment, stimulation of one such gamma axon also produced specific fusimotor effects on four afferents identified as coming from primary endings of muscle spindles. The gamma axon was therefore a fusimotor axon. The effect observed on stimulation of the gamma axon suggested a largely dynamic action. Other examples of gamma axons were encountered that on stimulation produced tension, but which could not be specifically associated with spindles. In addition, a number of gamma axons that did not develop tension were shown, on stimulation, to have fusimotor effects that were static in action. It is concluded that in extensively denervated muscles gamma motoneurones may sometimes sprout to innervate extrafusal fibres. The mechanical properties of the extrafusal fibres innervated by such gamma axons were similar to those of ordinary alpha motor units.


Respiratory network remains functional in a mature guinea pig brainstem isolated in vitro. We previously developed a perfused isolated brainstem preparation in the adult guinea pig (Morin-Surun and Denavit-Saubie 1989a) which permitted us to describe several types of rhythmic neuronal discharge. In the present study, we demonstrate that nearly all the periodic neuronal activity we recorded in the ventral respiratory areas were directly related to the respiratory-like periodic output of the hypoglossal nerve. This respiratory-like activity lasted several hours only when the brainstem was perfused by the basilar artery. This shows the necessity of the intraarterial perfusion to preserve a functional respiratory network. Analysis of the characteristics of hypoglossal respiratory-like activity shows that (1) two types of respiratory rhythms can be recorded; one with long respiratory phases (inspiratory and expiratory) and one with short respiratory phases. Depending on the preparation, either type occurs alone or intermingled with the other. (2) The shape of the inspiratory-like activity can change throughout the recording period while the periodicity remains stable. This preparation generates a respiratory rhythm and enables us to dissociate the different mechanisms involved in respiratory neurogenesis.


Corticofugal actions on lemniscal neurons of the cuneate, gracile and lateral cervical nuclei of the cat. Extracellular records were made from single identified lemniscal neurons of the cell-cluster regions of the cuneate and gracile nuclei, and of the lateral cervical nucleus, in pentobarbitone-anaesthetized cats. Forepaw, hind paw or face regions of the contralateral Sm I cortex were identified by recording through an inserted microelectrode which was then used for stimulation. The effect of a double cortical shock or train of shocks was usually inhibition: occasionally facilitation was observed, or mixed effects with facilitation preceding inhibition. Effects were seen in about half the cells studied in all three nuclei. Some cells of the lateral cervical nucleus were strongly excited, an effect not seen in the other nuclei. No component of these responses depended on suprathreshold stimulus intensities. Some lateral cervical cells were studied after deafferentiation by section of the dorsolateral spinal white matter; the same pattern of effects was seen. With an upper stimulus limit of 200 microA, cuneate but not gracile cells were affected from the cortical forepaw region, and gracile but not cuneate cells from the hind paw region. With threshold stimuli in an identified part of the forepaw cortical representation it was clear that cuneate cells with cutaneous receptive fields in corresponding parts of the forepaw had the lowest thresholds (minimum 6 microA). Threshold rose steeply with distance across the paw, suggesting quite sharp focusing of corticofugal effects in this system. When using similar procedures with the lateral cervical nucleus, with an upper limit of 200 microA, stimulation of forelimb cortex, or of facial cortex, affected both neurons with forelimb and those with hind limb fields.(ABSTRACT TRUNCATED AT 250 WORDS)


Development of human precision grip. II. Anticipatory control of isometric forces targeted for object's weight. The development of anticipatory control during lifts with the precision grip was examined in 100 children aged 1 to 15 years and in 15 adults. The children were instructed to lift an instrumented test object by using the precision grip between the thumb and index finger. The employed grip force, load force (vertical lifting force), vertical position and their corresponding time derivatives (i.e., grip and load force rates and acceleration) were recorded. The weight of the object was varied between trials to access the influence of the object's weight in the previous trial on the isometric force output. Already by the second year, children began to use information pertaining to the object's weight in the previous lift, i.e., they began to use an anticipatory control strategy. This occurred concomitant to the development of mainly bell shaped force rate profiles (Forssberg et al. 1991). The succeeding development of a more mature anticipatory control was gradual and adult-like capacity was not reached until 8-11 years of age.


Development of human precision grip. III. Integration of visual size cues during the programming of isometric forces. Recent evidence has shown that visual and haptical size information can be used by adults to estimate the weight of the object, forming the basis of the force programming during precision grip (Gordon et al. 1991a, b,). The present study examined the development of the capacity to use visual size information. In the first experiment, 30 children (age 1-7 years) and 10 adults performed a series of lifts with two boxes presented in an unpredictable order. The boxes were equal in weight but unequal in size and were attached to an instrumented grip handle which measured the employed grip force, load force, position and their corresponding time derivatives. The isometric force development was not influenced by the box size before the age of 3. However, the children aged 3 years and older demonstrated greater visual influences on the force programming than adults. To determine more precisely when children began to use visual size information, a second experiment in which the size and weight covaried was performed on 15 children. Children still did not use the size information during the force programming until the later half of the third year. It is concluded that this ability, probably involving associative transformations between the size and weight of objects, emerges around one year after anticipatory control based on somatosensory information pertaining to the weight of the object.


Phase-dependent reversal of reflexly induced movements during human gait. To investigate whether phase-dependent reversals in reflex responses on electromyography (EMG) are accompanied by movement reversals, a series of human volunteers were studied for their behavioural responses to sural nerve stimulation during running or walking on a treadmill. Low-intensity stimulation (less than 2.5 x perception threshold, T) of the sural nerve yielded facilitatory responses in the tibialis anterior muscle (TA), correlated with an induced ankle dorsiflexion (mean maximum 4 degrees) in early swing. The same stimuli yielded primarily TA suppression and weak ankle plantar flexion (mean maximum 1 degree) at end swing. The correlated induced knee angle changes did not precede the ankle changes, and they were relatively small. Mean maximum flexion in early swing was 6.2 degrees, while mean maximum extension was 3.7 degrees. High-intensity stimulation of the sural nerve (greater than 2.5 x T) always gave rise to suppression of the ongoing activity. This resulted in a second type of movement reversal. During late stance and early swing the responses in TA were suppressive (i.e. below background activity) and related to ankle plantar flexion. In contrast, the responses during early and middle stance consisted of suppression in extensor activity (gastrocnemius medialis and soleus) and ankle dorsiflexion. The data are discussed in terms of a new hypothesis, which states that the responses to electrical stimulation of cutaneous nerves during locomotion do not correspond directly to corrections for stumbling following mechanical perturbations during the step cycle. Instead, the data invite a reinterpretation in terms of the opening and closing of reflex pathways, presumably by a central pattern generator for locomotion.


The control of arm pointing movements in three dimensions. In this study we investigated pointing movements made with an extended arm. Despite the large number of mechanical degrees of freedom, limb orientation adopted during pointing could be described by rotation axes contained on a two-dimensional curved surface. As a result of the curvature, the orientation of a linear plane approximating a small region of the curved surface was dependent on the location of the movements within the full workspace. These results account for earlier suggestions that limb orientation could be described by coplanar rotation vectors and that the orientation of the plane moved with the workspace. Despite the additional complexity, our results indicate that the number of degrees of freedom used to position the extended forearm is reduced from four to two for normal pointing movements. Contributions to orientation of the wrist and hand by supination/pronation of the forearm were minor for changes in shoulder yaw angle. However, supination/pronation added significantly to orientation of the hand for changes in shoulder pitch angle.


Motor strategies in landing from a jump: the role of skill in task execution. A motor performance which involves multijoint coordination and belongs to the natural repertoire of motor behavior has been studied. Displacements have been related to EMG in the lower limb when taking off and landing from a jump down (45 cm) onto two surfaces of differing compliance in two populations of teenage girls: skilled and unskilled. To evaluate the performance, an index was defined taking into account: 1) the time required for reaching stability (1 body weight) after landing, and 2) the amount of sway during the stabilization time. Despite the apparent intra and inter subject similarities in performing the jump-down, slight differences were observed in both the kinematics and electromyogram patterns. During takeoff, two strategies were identified that were not related to either skill or landing surface compliance. The most common strategy, "Push Off", is characterized by almost full joint extension when departing from the jump platform and includes a swing period during flight. The other strategy, "Roll Off", is characterized by joint flexion at departure and continual extension during midflight. While the ankle dorsiflexor, tibialis anterior, is active in preparation for the takeoff phase in both strategies, it is followed by activation of the ankle plantarflexors, lateral gastrocnemious and soleus and the hip/knee musculature, rectus femoris, biceps femoris, and vastus lateralis, only in the push off strategy. The roll off strategy is characterized by a lack of other muscle activation prior to takeoff. At landing, regardless of the strategy used in takeoff, onset of muscles followed the same sequence for both landing surfaces; ankle musculature activity began first followed by activity in the knee and hip musculature. The onset of the musculature occurred closer to landing when landing on the more compliant surface. Skilled subjects were characterized by adjustments in amount of ankle extension present at landing and concomitant flexion post-landing with respect to landing surface. When landing on the rigid surface, the ankle was more plantarflexed and onset of the dorsiflexor occurred after that of the plantarflexors; on foam, dorsiflexor activity was coincident with the plantarflexors. Ankle joint range of motion post-landing was subsequently larger when landing on the rigid surface. In contrast, unskilled individuals used a default strategy for landing on both surfaces where the ankle position and movement was between that seen for the two conditions in the skilled individual. It is suggested the landing and takeoff phases are programmed independently in both skilled and unskilled subjects.(ABSTRACT TRUNCATED AT 400 WORDS)


cFos labeling in rat superior colliculus: activation by normal retinal pathways and pathways from intracranial retinal transplants. Previous studies in this laboratory have shown that intracranial retinal transplants can establish both anatomical and functional connections with the host brain. Embryonic rat retinae transplanted intracranially into neonatal host brains are capable of evoking appropriate physiological and behavioral responses when illuminated. The present study employs the specific expression of the cFos protein to identify brain regions in which immediate-early gene activation can be recognized in response to flash stimuli delivered to retinal transplants and to normal intact retinae. Stimulation of the intact eye induced significant cFos expression in various visual centers, including the stratum griseum superficiale of the superior colliculus and the pretectal area, but not in the dLGN, suprachiasmatic nucleus, or retina. Animals with functional transplants expressed cFos throughout the depth of the stratum griseum superficiale and stratum opticum of the superior colliculus, thus apparently activating an additional population of superior collicular cells. Like the eye-stimulated animals, animals with functional transplants failed to elicit significant cFos expression in the dLGN or transplanted retina. This study indicates that intracranial retinal transplants are capable of forming functional connections with the host superior colliculus which not only mediate transient changes in electrical activity but also may affect gene expression through the induction of the c-fos gene.


Immunohistochemical changes of neuronal calcium-binding proteins parvalbumin and calbindin-D-28k following unilateral deafferentation in the rat visual system. The neuron-specific calcium-binding proteins, parvalbumin and calbindin-D-28k, were studied in the subcortical visual system of normal and unilaterally deafferented albino rats. Immunohistochemistry with monoclonal antibodies was used on vibratome sections through optic tract (OT), dorsal lateral geniculate nucleus (dLGN), olivary pretectal nucleus (OPN), and superior colliculus (SC). In controls, OT stained strongly for parvalbumin and weakly for calbindin-D-28k. The dLGN contained a plexus of parvalbumin-positive fibers. In dLGN, calbindin-D-28k-antibodies showed strong labeling of some neurons with long dendrites and weak staining of the cytoplasm in other neurons. In OPN, parvalbumin stained a ring of neurons and terminals in the shell region, whereas calbindin-D-28k was contained in medial cell populations. In SC, parvalbumin was contained in fibers, terminals, and neurons throughout the visual layer. Calbindin-D-28k showed a laminar distribution of neurons with a predominance in deep portions of superficial grey matter and in ventral portions of stratum opticum. Following unilateral deafferentation induced by optic nerve section, retinal axons showed immunohistochemical changes related to Wallerian degeneration and target neurons reacted by changes of calcium-binding proteins. Parvalbumin and calbindin-D-28k immunostaining decreased during Wallerian degeneration of OT. In the deafferented dLGN, immunohistochemical labeling for calbindin-D-28k declined in strongly stained neurons from 4 to 21 days after lesion. Measurement of dendritic length per number of cells or per area of dLGN showed a significant decline for the contralateral side at 4, 8, and 21 days (ANOVA, P less than 0.05). In deafferented OPN, terminal-like staining for parvalbumin decreased and neuronal labeling was enhanced. In deafferented SC, the neuronal and dendritic staining for parvalbumin increased beginning from Day 1 on and persisting at Day 21, whereas fibers and terminal-like elements decreased in staining. Measurement of parvalbumin-positive neurons per area of SC showed a significant increase of labeling in the contralateral side from Day 1 to Day 21 (ANOVA, P less than 0.05). These studies show that cellular responses to deafferentation of visual neurons involve a regulation of calcium-binding proteins. The decline in staining for calbindin-D-28k in dLGN may relate to reduced retinal afferent activity. The progressive cellular changes in parvalbumin staining may be related to unmasking of intrinsic neurons after removal of parvalbumin-containing, afferent fibers and terminals. Additionally, the changes of parvalbumin labeling in SC neurons may reflect a plastic reorganization of local circuits known to occur in rat SC in response to deafferentation.


Observation of the blood-brain barrier function and vasomotor response in rat microcirculation using intravital fluorescence microscopy. The blood-brain barrier function and the vasomotor response in rat microcirculation was studied using a closed cranial window technique and an intravital fluorescence microscope with an attempt to reduce both the light intensity and the amount of 2% Na(+)-fluorescein to as little as possible. The light intensities in the focus were less than or equal to 2.5 mW/cm2, and with the low-light TV camera we were able to reduce the total amount of the dye to less than or equal to 0.7 ml during 6-h experiments. Observing the brain surface every 30 min showed that the barrier function remained intact for up to 6.0 h following the first iv administration of the dye. This study has reconfirmed that the pial vessels have physiological vasomotor responses (e.g., CO2 reactivity). The osmotic barrier disruption started at a small venule 10 min after the start of superfusion with mannitol (1000 mOsm/liter). In conclusion, the microscope system and the small animal model used in this study were useful in studying the blood-brain barrier function and the vasomotor response simultaneously under various experimental conditions.


Alteration of second-messenger ligand binding following 2-hr hemispheric ischemia in the gerbil brain. The alterations of second-messenger ligand binding and cerebral blood flow (CBF) were evaluated in the gerbil brain after 2-h unilateral common carotid artery occlusion. [3H]Forskolin (FK) and [3H]phorbol-12,13-dibutyrate (PDBu) were used as specific ligands for adenylate cyclase (AC) and protein kinase C (PKC) activity estimation, respectively. CBF was determined at the end of the experiment by the [14C]iodoantipyrine method. A quantitative autoradiographic method permitted simultaneous measurement of the three parameters in the same brain. The levels in the caudate-putamen, globus pallidus, and hippocampus were analyzed. The animals were divided into three groups: Group 1 with severe ischemia (CBF in the lateral nuclei of the thalamus (CBFt) less than 50 ml/100 g/min), Group 2 with mild ischemia (CBFt greater than or equal to 50 ml/100 g/min), and the Sham Group. The PDBu binding revealed a statistically significant increase in the caudate-putamen, lateral nuclei of the thalamus and hippocampus (CA1 and CA3 regions and dentate gyrus) on the ischemic side in Group 1 as compared to that in Group 2 and the Sham Group. In contrast, the FK binding did not show any significant changes in any of the regions. These data and our previous findings for 6-h ischemia suggest that (1) PKC translocation to the cell membrane may occur at the early ischemic phase in particular regions including the caudate-putamen, lateral nuclei of the thalamus and hippocampus, with the translocated PKC gradually diminishing during the subsequent ischemic period; and (2) the suppression of the AC system observed in 6-h ischemia may not appear in the early ischemic phase.


Extracellular proteins of goldfish optic tectum labeled by intraocular injection of 3H-proline. A prominent group of soluble glycoproteins with a molecular weight of 30K-40K and pI 5.0-5.6 was detected in various parts of the goldfish brain as well as in the optic nerves. Since these proteins are readily liberated from the tissue, we have designated them exoglycoproteins (X-GPs). The X-GPs in the optic tectum were found to be labeled after intraocular injection of radioactive tracers, in a manner consistent with the labeling of proteins transported in the optic axons. However, the labeling of X-GPs was blocked by intracranial injection of a protein synthesis inhibitor, whereas the labeling of axonally transported proteins was unaffected. This shows that the X-GPs can be synthesized locally within the brain. Nevertheless, when protein synthesis in the retina was blocked, the labeling of the X-GPs in the tectum was prevented, like the labeling of axonally transported proteins. Thus precursors for the synthesis of X-GPs can be derived from materials transported in the optic axons. This synthesis can occur in nonneuronal cells, as indicated by the incorporation of labeled amino acid into X-GPs in optic nerves directly exposed to the label. The synthesis of X-GPs was increased in regenerating nerves, suggesting that these proteins may play a role in regeneration. Partial amino acid sequencing of the proteins showed that they are identical to the proteins previously identified as "ependymins," which have been implicated in neuronal plasticity. There are minor differences in amino acid sequence among some individual spots.


Quisqualic acid-induced lesion of the nucleus basalis of Meynert in young and aging rats: plasticity of surviving NGF receptor-positive cholinergic neurons. Previous studies have demonstrated that cholinergic neurons in the adult rat forebrain, i.e., septal region, are able to respond and regenerate after damage followed by exogenous treatment with beta-nerve growth factor. Furthermore, it has been shown that an age-related loss of NGF-receptor (NGFr) immunoreactivity occurs in cholinergic septal neurons. Since the regenerative capacity of cholinergic neurons is of importance for potential therapeutic strategies during the course of age-related neurodegenerative diseases, we have compared NGFr positive neurons in young adult (3 months old) and in aging (18-24 months old) rats in their ability to produce a physiological plasticity response after surviving an excitotoxic lesion of the nucleus basalis of Meynert (NBM). In aging control rats, NGFr immunoreactivity within NBM neurons was significantly reduced, in analogy to data obtained earlier from studies about septal neurons in aged rats. After lesion with quisqualic acid, a severe cell loss as well as atrophy of remaining cholinergic neurons was observed in both groups. Investigation of the NBM at various times after the lesion demonstrated signs of axonal or dendritic sprouting and local regeneration, with a maximum seen 3 months after the lesion. No age-related differences in the response could be found. However, despite local fiber growth, no reinnervation of the frontal and parietal cortex could be noted, as demonstrated by acetylcholinesterase histochemistry. Our findings suggest that, despite a relatively early onset of NGFr decline during lifetime, cholinergic cells keep the capacity for a plastic response, although they ultimately fail to reinnervate the neocortex.(ABSTRACT TRUNCATED AT 250 WORDS)


Spreading depression induces prolonged reduction of cortical blood flow reactivity in the rat. The purpose of the present study was to examine the dynamic aspects of the cerebrovascular events occurring during and up to 2 h following cortical spreading depression (CSD) in the rat, using the mass spectrometry technique which enables continuous measurements of the cortical tissue PO2 and PCO2 and repeated blood flow measurements (CoBF) by helium clearance. We mostly sought to determine whether cortical perforation by a stimulation electrode induced long-lasting perturbation of the cortical vasoreactivity to hypercapnia and basal forebrain electrical stimulation. Cortical perforation in the animal under alpha-chloralose anesthesia, chronically implanted with mass spectrometry probes, was associated with biphasic changes in tissue gases. PO2 first briefly decreased (-7.8%) and then strongly increased (+79%) while PCO2 changed in the opposite direction (+7%, -13%) in the ipsilateral frontal cortex. Qualitatively similar changes occurred in the ipsilateral parietal cortex. The CoBF measurements showed a marked vasodilation (131 and 108% in the frontal and parietal cortex, respectively) in parallel with the PO2 increase, followed by a prolonged (60 min), moderate hypoperfusion (maximum -17% at 20 min after CSD). There was a pronounced reduction of vascular reactivity to both hypercapnia (20.3% of the control response) and substantia innominata stimulation (1/6 of the response obtained 80 min later) at 10 min after CSD. Both reactivities progressively recovered in approximately 2 h. Since the issue of CSD in human has become a popular hypothesis for migraine, the reduced cerebrovascular reactivity could constitute the basis of a test for the involvement of CSD in migraine.


Forelimb motor performance following cervical spinal cord contusion injury in the rat. The purpose of this study was to examine the degree, persistence, and nature of forelimb behavioral deficits following cervical spinal cord contusion injury in the rat. Forelimb reaching and pellet retrieval, forehead adhesive sticker removal, and vibrissae-induced forelimb placing were examined for 16 weeks following a weight-drop injury (10.0 g-2.5 cm) at the C4-C5 spinal level. Nine of 13 rats studied were unable to perform the pellet retrieval task due to pronounced forelimb extension hypometria. However, these animals did carry out the forehead sticker removal and vibrissae-induced placing tasks. Therefore, the loss of reaching ability related to pellet retrieval was not due to generalized paralysis. This interpretation was further supported by evaluation of the rostrocaudal extent of relative motoneuron loss from 1-mm divisions through the lesion zone. The extent of motoneuron pathology ranged from 2 to 6 mm but was largely confined to the C4-C5 spinal segments. Morphometric assessments of axonal sparing revealed that pellet retrieval performance during the last month of observation was significantly correlated with fiber sparing in the dorsal columns and ventral white matter, whereas no significant correlation could be demonstrated with regard to dorsolateral white matter. While there were no conspicuous differences in qualitative assessments of damage to interneuron pools (i.e., laminae V to VII) between the nonreaching and retrieval-recovered rats, the possibility of combined white and gray matter pathology contributing to this deficit still exists. These initial findings thus demonstrate that the weight-drop contusion injury model can be adopted to studies of cervical spinal cord trauma in the rat. Such lesions yield permanent deficits in forelimb function lending to future studies of possible therapeutic interventions. Furthermore, performance deficits observed at 1 week postinjury in the placing and forehead sticker removal tasks can be predictive of any potential for long-range spontaneous recovery in pellet retrieval ability.


Axonal conduction and electrical coupling in regenerating earthworm giant axons. Severed halves of medial giant axons (MGAs) and lateral giant axons (LGAs) in earthworms survive and are functionally reconnected as early as the first postoperative week. During the first 150 postoperative days, there is an increase in conduction velocity of action potentials and strength of electrotonic coupling between the severed axonal stumps across the lesion site. Electrophysiological analyses suggest that this functional reconnection occurs by transmission of action potentials through the lesion site by active propagation along neurites which make electrotonic connections rather than chemical synapses. The regenerated connections restore the original connectivity pattern for conduction of action potentials or spread of electrotonic potentials; i.e., MGA stumps reconnect with MGA stumps, and LGA stumps with LGA stumps. These and other data suggest that the mechanisms responsible for establishing appropriate functional reconnection of severed earthworm giant axons requires cell-specific matching of axons and neurites, rather than a competition between appropriate and inappropriate functional connections.


Restoration of orbicularis oculi function by contralateral orbicularis oculi innervated muscle flap vs neuromuscular pedicle technique. In preliminary experiments with dogs and cats, unilateral paralysis of the orbicularis oculi muscle group was produced by a section of the seventh nerve that included the posterior auricular branch. Either one of two procedures was then employed in attempts to reinnervate the paralyzed eyelid. In one group of animals, a neuromuscular pedicle was employed and in another, a contralateral orbicularis innervated muscle flap was used. Both methods restored synchronous, reflex blinking to the denervated eyelid. Of the two procedures, neurotization appears to offer the greater promise because the use of a neuromuscular pedicle requires an expendable nerve that is functional, and no such suitable substitute is available in humans.


Long-latency auditory-evoked potentials: role of polysensory association cortex in the cat. The objective of this study has been to define the role of polysensory association cortex in the generation of "wave NA" and of "wave C," long-latency auditory-evoked potentials recorded from the vertex of conscious cats as, respectively, a marked negative potential of latency 30-48 msec followed by a broad positive wave of latency 50-75 msec. Wave C may represent the feline analogue of the longer latency human auditory-evoked potential wave P2, insofar as both waveforms are very large amplitude, long duration positivities characterized by long recovery cycles. Based on previous studies of wave C and the generators of other middle-latency evoked potentials, we hypothesized that both wave NA and wave C might reflect, at least in part, the cortical culmination of a nonlemniscal line auditory association system arising in reticulothalamic projections to intralaminar and associated ventral thalamic regions. Relays from these thalamic areas are known to project to polysensory association cortex, including pericruciate gyrus, anterolateral gyrus, and medial suprasylvian gyrus. Therefore we implemented a series of lesion experiments to characterize the role of each of these cortical areas in the production of wave NA and wave C. Our results indicate that all three polysensory association areas contribute significantly to both waves NA and C, although the largest effects followed ablation of the pericruciate area alone. Thus, the generator substrates of waves NA and C appear to involve a long-recovery cycle system which functionally incorporates activation of association cortex.


Threshold relationship between cerebral blood flow, glucose utilization, and energy metabolites during development of stroke in gerbils. Focal brain ischemia was produced in halothane-anesthetized Mongolian gerbils by occluding the right common and the left external carotid artery. Ninety minutes after vascular occlusion the following regional hemodynamic and metabolic parameters were evaluated in adjacent cryostat sections taken from seven different coronal planes of each brain: cerebral blood flow (CBF), glucose utilization (CMRG), and the tissue content of ATP and glucose. NADH fluorescence was recorded from the surface of the cryostat block. In addition, tissue slices were taken from each brain to determine the rate of phosphorylation of 2-deoxyglucose in ischemic and nonischemic regions. Depending on the density of ischemia, the following metabolic disturbances were observed. At CBF values below 35 ml x 100 g-1 x min-1 CMRG increased and at values below 25 ml x 100 g-1 x min-1 it declined sharply. Glucose content declined when CBF was below 35 ml x 100 g-1 x min-1 and ATP fell at CBF below 20 ml x 100 g-1 x min-1. At 10 ml x 100 g-1 x min-1 ATP was completely depleted. NADH fluorescence was found elevated at flow rates that caused an increase of glucose utilization and was maximal when CBF stopped. The ischemic thresholds for the initial increase in CMRG and the complete depletion of ATP content represent the metabolic equivalent of the penumbra zone and provide a basis for the evaluation of therapeutic procedures for the treatment of stroke.


MR imaging for measurements of ventricles and cerebral cortex in postnatal rats (H-Tx strain) with progressive inherited hydrocephalus. As part of a wider investigation on brain structure and function in young rats with inherited hydrocephalus, this study was undertaken to develop a technique to monitor, and quantify, progressive changes in ventricle and cortical dimensions in hydrocephalic rats in vivo. Magnetic resonance images were obtained with a T1-weighted protocol and a multislice variant of the inversion recovery sequence which displayed optimal contrast between cerebrospinal fluid and brain parenchyma. Rats were imaged at 7, 14, and 21 days after birth by taking 10 contiguous 2.0-mm coronal slices. Area and distance measurements were taken from the displayed images, enabling calculation of ventricle volumes, cortical length, and cortical thickness. The lateral ventricle volume, which was already 100-fold larger than control at 7 days, increased to over 1000 mm3 at 21 days. The anterior-posterior length of the cortex was up to 27% larger in hydrocephalics and the cortical mantle was up to 75% thinner, particularly in the posterior cortex. Subsequent fixation and histological sectioning of two 21-day-old rats showed that the smaller dimensions, which were obtained from analysis of the sections, could be accounted for by shrinkage of the tissue during dehydration and wax embedding.


Disappearance of astrocytes and invasion of macrophages following crush injury of adult rodent optic nerves: implications for regeneration. Injury to the mammalian central nervous system results in loss of function because of its inability to regenerate. It has been postulated that some axons in the mammalian central nervous system have the ability to regenerate but fail to do so because of the inhospitable nature of surrounding glial cells. For example, mature oligodendrocytes were shown to inhibit axonal growth, and astrocytes were shown to form scar tissue that is nonsupportive for growth. In the present study we report an additional phenomenon which might explain the failure of axons to elongate across the site of the injury, namely, the absence of astrocytes from the crush site between the glial scar and the distal stump. Astrocytes began to disappear from the injury site as early as 2 days after the injury. After 1 week the site was necrotic and contained very few glial cells and numerous macrophages. Disappearance of glial cells was demonstrated in both rabbit and rat optic nerves by light microscopy, using antibodies directed against glial fibrillary acidic protein, and by transmission electron microscopy. Results are discussed with reference to possible implications of the long-lasting absence of astrocytes from the injury site, especially in view of the differences between the present findings in rodents and our recent observations in fish.


Magnetic resonance imaging to monitor pathology of caudate-putamen after excitotoxin-induced neuronal loss in the nonhuman primate brain. We used MR imaging to locate and monitor in vivo the pathological events taking place 2 to 4 weeks after unilateral striatal injections of ibotenic acid (IA) in the Papio papio baboon. As early as 2 weeks after IA injections, excitotoxic lesions in the caudate and the putamen were directly visualized on T1-weighted images as small areas of low signal intensity. On T2-weighted images, the lesion sites were visualized as areas of high-intensity signal, spreading over larger areas than the corresponding regions in T1-weighted images. These alterations of T2-values in the lesioned striatum persisted 4 weeks after surgery. However, as the striatal degeneration progressed from 2 to 4 weeks after lesion, the size of the areas of high signal intensity on T2-weighted images decreased, whereas the same regions appeared essentially unmodified on T1-weighted images. A marked enlargement of the ipsilateral lateral ventricle (a characteristic of excitotoxic striatal lesions) could be detected 4 weeks after surgery, on both axial T1- and T2-weighted images. Comparisons of MR images with postmortem anatomical data indicated that areas of increased T1 values corresponded to regions of severe neuronal depletion (a direct result of the excitotoxic lesion), whereas areas of increased T2 values corresponded to regions of increased content in astrocytes and ferritin and probably in the early period following lesion (2 weeks) to a superimposed edema.


Effects of adrenal medulla grafts on plasma catecholamines and rotational behavior. The mechanisms by which adrenal medulla grafts influence the function of host brains in animal models of Parkinson's disease are unclear. To explore this issue, fragments of adrenal medulla or sciatic nerve were transplanted into the lateral ventricle of bilaterally adrenalectomized (ADX) or sham-ADX rats with unilateral 6-hydroxydopamine lesions of the substantia nigra. Additional control group received sham-transplantation surgery. Behavioral effects of these procedures were tested following administration of apomorphine, amphetamine, or nicotine. Plasma catecholamines were measured before and after transplantation surgery. In both ADX and sham-ADX rats, adrenal medulla grafts produced greater decreases in apomorphine-induced rotational behavior than did sciatic nerve grafts or sham-transplanted groups. Decreases in rotation were smaller in ADX than in sham-ADX animals, regardless of graft treatment. Plasma catecholamines increased after transplantation surgery in each of the sham-ADX groups, regardless of graft type. Increases in plasma dopamine concentrations were associated with decreases in rotational behavior. Five months after transplantation, grafted chromaffin cells demonstrated catecholamine fluorescence, tyrosine hydroxylase (TH) and chromogranin A immunoreactivities, and expression of TH mRNA. It is concluded that adrenal medulla grafts produce decreases in apomorphine-induced rotation through a combination of two independent effects. One is a specific effect of adrenal medulla grafts. The second is a nonspecific effect that requires an intact adrenal gland and may be related to increases in plasma catecholamine concentrations.


Basic fibroblast growth factor prevents neuronal death and atrophy of retrogradely labeled preganglionic neurons in vivo. We have used retrograde fluorescent tracing (fast blue) both before (prelabeling) and at time points after selective unilateral adrenomedullectomy in vivo (post-labeling) in order to investigate the survival and morphology of sympathoadrenal preganglionic (SAP) neurons located in the lower thoracic intermediolateral (IML) cell column of the adult rat spinal cord. By prelabeling with fast blue it was found that the majority (i.e., more than 85%) of the SAP neurons underwent degeneration and were lost from the IML cell column within 4 weeks after peripheral target lesion and administration of intramedullary gelfoam implants containing a nontrophic control protein cytochrome c. By contrast, atrophy and loss of SAP neurons was largely prevented by local treatment (intramedullary implants) with recombinant basic fibroblast growth factor (bFGF) as determined from counts of large and "healthy-looking" prelabeled neurons within the IML column ipsilateral to the lesioned (i.e., operated) side 4 weeks postimplantation. The time course of withdrawal of preganglionic axons from their lesioned target area was investigated by fast blue injections into intramedullary (control or bFGF) implants at weekly intervals postimplantation (postlabeling) and was documented by counting the number of healthy SAP neurons that retained the label. Without bFGF treatment, progressive numerical loss of SAP neurons was evident within 1 to 4 weeks postlesioning, indicative of pronounced retrograde cell degeneration. Retrograde cell degeneration was insignificant during the first 2 weeks postimplantation after early postlesion treatment with exogenous bFGF; it was apparently postponed to occur after 1 month. Implantation of gelfoam containing neutralizing anti-bFGF-antibodies resulted in accelerated retrograde axon degeneration implying that bFGF is an endogenous trophic factor for SAP neurons. The results are consistent with the idea that SAP neurons actually die following peripheral target lesion and are not supported from other trophic sources. However, these neurons can be prevented from disconnection-induced death by providing exogenous bFGF. Limited amounts of endogenous FGF may also become available to SAP axons by disintegration of nerve terminal-surrounding cells delaying the process of retrograde SAP neuron death.


Neurotrophic effects of steroids on lesion-induced growth in the hippocampus. II. Hormone replacement. The mediation of lesion-induced sprouting in the nervous system is a complex interaction of local membrane factors and circulating hormones. This series of studies examines the reactivity of the sprouting response of both male and female subjects under different hormonal conditions. Young adult male and female Sprague-Dawley rats which were gonadectomized (GDX) and adrenalectomized (ADX) underwent a unilateral entorhinal cortex lesion, which partially denervates the molecular layer of the ipsilateral hippocampal denate gyrus. At the time of the lesion, each animal received hormonal therapy. Fifteen days post-ERC-ablation the brains were analyzed for changes in reactive fiber outgrowth in the hippocampal commissural/associational afferents. Fiber outgrowth in females in the "asteroidal" (GDX/ADX) condition was unaffected. Asteroidal males demonstrated a decreased response. Gonadal steroid replacement, estrogen or testosterone, enhanced outgrowth in both asteroidal males and females. Glucocorticoid replacement suppressed outgrowth in both asteroidal males and females. Gonadal steroids clearly have neurotrophic activity which is interactive with glucocorticoid activity. Glucocorticoids under the GDX/ADX conditions in vivo have a negative impact on fiber outgrowth in both sexes. The effect of glucocorticoids is most dramatic when compared to the outgrowth of asteroidal animals without additional hormonal supplementation.


Myelinated fiber regeneration after sciatic nerve crush: morphometric observations in young adult and aging mice and the effects of macrophage suppression and conditioning lesions. To study myelinated nerve fiber regeneration during aging, the right sciatic nerves of 6- and 24-month-old mice were crushed at the sciatic notch. Two, 4, and 8 weeks later, both groups of mice were perfused. The sciatic nerves were processed so that the transverse sections of each nerve subsequently studied by light and electron microscopy included the entire posterior tibial fascicle 5 mm distal to the crush site. Two weeks after axotomy, fascicles of aging mice contained significantly fewer regenerated myelinated fibers than those of young adults. After 4 weeks, the difference in the number of myelinated fibers was less. However, measurements of myelinated fibers in fascicles of aging mice showed that areas of Schwann cell cytoplasm and myelin were significantly reduced at all intervals. In contrast, although axon diameters in aging mice were somewhat less 2 weeks after crushing, the difference decreased with time, suggesting that in nerves of aging mice, regenerative responses of Schwann cells were more affected than those of axons. Other experiments in young mice showed that myelinated fiber regeneration could be retarded by suppressing macrophage responses and was not significantly changed by conditioning lesions before crush injury.


Divergent effects of astroglial and microglial secretions on neuron growth and survival. Brain glia have a secretory capacity which can modulate neuronal function. Astrocytes release proteins which enhance neuronal survival and induce neuronal growth and differentiation. These effects can be blocked by antagonists of voltage-dependent calcium channels and may be partly mimicked by Bay K 8644, a calcium channel agonist. Two of these neurotrophic proteins appear, on the basis of their physical properties and effects on ciliary ganglion neurons, to be ciliary neurotrophic factor and basic fibroblast growth factor. Activated microglia release a heat- and protease-stable neurotoxin of low molecular weight. This neurotoxicity is blocked by NMDA receptor antagonists. Ciliary neurons exposed to the microglial neurotoxin exhibit an abnormal distribution of neurofilament immunoreactivity, which becomes concentrated in a perinuclear region, while the astroglial growth factors induce neurofilament organization into an extensive neuritic network. The astrocyte-released growth factors can counteract the effect of the microglial neurotoxin and lead to unimpaired neural differentiation in the presence of the neurotoxin.


Human fetal xenografts of brainstem tissue containing locus coeruleus neurons: functional and structural studies of intraocular grafts in athymic nude rats. Fetal human brainstem tissue including the nucleus locus coeruleus was transplanted to the anterior eye chamber of athymic nude rats. Most transplants survived and grew in the anterior chamber of the eye. After 9-15 months, the host animals were anesthetized and electrophysiological or in vivo electrochemical recordings were performed. The brainstem transplants contained spontaneously active neurons with regular single-spike firing patterns. The neurons responded to ipsilateral light stimulation with an increase in firing rate and to the alpha 2-receptor agonist clonidine with significantly decreased firing rates. In vivo electrochemical studies demonstrated reproducible noradrenergic overflow after local application of potassium. Immunohistochemical evaluation of the brainstem transplants showed an abundance of tyrosine hydroxylase-positive neurons and neurites in all transplants and a dense network of neurofilament-, synapsin-, and glial fibrillary acidic protein-positive profiles throughout the grafts. Taken together, the present physiological and histochemical data indicate that it is possible to obtain transplants containing a specific monoaminergic population within the brainstem from human fetal fragments and to maintain these transplants in oculo in athymic nude rats for at least 15 months, during which time noradrenergic neurons develop.


Transynaptic regulation of low-affinity p75 nerve growth factor receptor mRNA precedes and accompanies lesion-induced collateral neuronal sprouting. The bilateral sympathetic innervation of the rat pineal gland from the two superior cervical ganglia (SCG) is a useful model system to investigate the mechanisms by which intact neurons compensate for neuronal losses. Cutting of the internal carotid nerve (ICN) on one side has been shown to result in the removal of approximately one-half of the innervation to the pineal gland within 2 days. This denervation is followed by the development of collateral neuronal sprouting from the contralateral "intact" SCG, most of which takes place during the next 2 days. Using a solution hybridization protection assay, levels of low-affinity NGF receptor p75NGFR mRNA (pg/microgram total RNA) were found to be increased 25%, with no change in cyclophilin mRNA, in the SCG contralateral to the lesion performed 1 or 3 days earlier. In situ hybridization with a 35S riboprobe complementary to p75NGFR mRNA demonstrated a large increase in this mRNA in some cells of this intact SCG at both 1 and 3 days after a contralateral ICN cut lesion. The clustering of these cells toward the rostral portion of the SCG suggests that they may overlap with the population of sympathetic neurons which provides innervation to bilaterally innervated structures such as the pineal gland. The nature of the signals involved in the regulation of NGF receptor mRNA levels and their role in initiating and maintaining collateral sprouting remain to be fully established. Nevertheless, the time course of the changes in mRNA levels suggests that regulation of the low-affinity NGF receptor gene may be involved in the sequence of events associated with the collateral sprouting response by intact sympathetic nerve cells following partial denervation.


Sensory nerves impair sympathetic reinnervation and recovery of smooth muscle function. Neuronal populations projecting to a common target may compete for neurotrophic substances. To determine if competition impairs target reinnervation, we examined the effect of capsaicin-induced sensory denervation on sympathetic nerve ingrowth to the sympathectomized rat superior tarsal smooth muscle. In tarsal muscles with intact sympathetic innervation, capsaicin injection on Day 2 reduced numbers of perimuscular CGRP-ir sensory nerves by 68% at 3-4 months; however, it did not alter dopamine-beta-hydroxylase-ir nerve density, response to nerve stimulation, or tarsal muscle adrenoceptor-mediated contraction. Tarsal muscles denervated by ipsilateral superior cervical ganglionectomy on Postnatal Day 4 were partially reinnervated by fibers from the contralateral ganglion, as noted in previous studies. Sensory denervation by capsaicin improved sympathetic reinnervation, as evidenced by a 174% increase in numbers of DBH-ir nerves and a 62% increase in neurally mediated smooth muscle contraction evoked by electrical stimulation of the contralateral pathway relative to reinnervated muscles of vehicle-injected rats; smooth muscle function was also influenced, as indicated by a decrease toward normal in adrenoceptor sensitivity. Tarsal muscles denervated at 30 days were not reinnervated in either vehicle-injected or capsaicin-treated rats, indicating that sensory denervation does not extend the developmental window during which contralateral reinnervation can occur. Both the vehicle-injected and capsaicin-treated preparations with sustained juvenile sympathectomy showed sensory hyperinnervation as adults; thus, a chronic reduction in competition from sympathetics is a sufficiently powerful stimulus to overcome the decreased nerve density induced by neonatal capsaicin treatment. We conclude that sensory nerves limit the extent of sympathetic reinnervation and functional recovery that can occur following neonatal sympathetic denervation.


Abnormal cerebellar output in rats with an inherited movement disorder. Biochemical and metabolic mapping techniques have consistently identified the deep cerebellar nuclei (DCN) of the genetically dystonic rat as a site of abnormality. Extracellular single-unit recording techniques were used to assess the functional significance of these findings in affected rats and normal littermates between 16 and 25 days of age. Cells in the medial nucleus of the mutant rats had significantly increased spontaneous firing rates in comparison with cells from normal rats. In both the medial and the interpositus nuclei, cells from the mutants fired more rhythmically than those from the normal rats. When harmaline was administered systemically to activate the olivo-cerebellar system, in normal rats, increased firing rate and bursting patterns of activity were seen. There was no reliable change in the average firing rate or rhythmicity of cells in the medial nucleus of the dystonic rats, although previous studies have shown that harmaline activates neurons in the inferior olive in the mutants. It is likely that naturally stimulated olivary activity also fails to modulate cerebellar output in this model of inherited movement disorder. Anatomical studies did not reveal any consistent changes in the number of Purkinje cells, the volume of the DCN, or the soma size of DCN neurons. Since the electrophysiological findings cannot be ascribed to a loss of the Purkinje cells that normally provide an inhibitory input to the cerebellar nuclei, the results of this study indicate the presence of a functional defect in the control of cerebellar output in the dystonic rat that accounts for the failure of these animals to display harmaline tremor and which may be critical to the motor syndrome.


Mechanisms of circulatory homeostasis and response in Aplysia. This review concerns the organization and function of arterial vasculature in Aplysia californica, especially the vasomotor reflexes that support circulatory homeostasis, and fixed patterns of response that may reroute blood flow during changes in behavioral state. The observations presented here raise three hypotheses for further study: 1) Arterial vasculature is functionally organized with precisely structured, independently regulated subdivisions; these are most evident for arterial systems serving digestive and reproductive processes; 2) arterial musculature is inherently responsive to local pressure changes, having both static and dynamic reflexes that promote efficient, evenly-distributed flow of blood; and 3) complex, long-lasting behaviors like egg laying have, as part of their makeup, equally prolonged and stereotypical changes in the pattern of circulation. Taken together, these observations support the view that maintenance and adjustment of blood flow in gastropod molluscs is an unexpectedly complex and highly integrated component of behavior.


Evidence for the induction and release of pancreatic folylpolyglutamate hydrolase by the ingestion of folyl polyglutamates in the rat. Pteroyl polyglutamate hydrolase (folyl conjugase), which hydrolyses the dietary polyglutamyl folates into simple forms prior to absorption, has been shown to be induced in rat pancreas in response to dietary polyglutamyl folates but not by ingestion of synthetic unconjugated folates. Folate absorption, as measured by the rise in serum folate levels after ingestion, of dietary conjugated folates is impaired in pancreatectomised animals whereas absorption of synthetic simple folate is not. A severe build-up of folyl conjugase is observed in the lumen of control but not in pancreatectomised animals after dietary folate ingestion. These results taken together would suggest that dietary folates are hydrolysed to monoglutamyl forms suitable for absorption in the lumen; the hydrolysis is catalysed by a luminal folyl conjugase of pancreatic origin induced by dietary conjugated folates.


Short cerebral ischemia and subsequent reperfusion and treatment with stobadine. Lipid peroxidation and activities of antioxidative enzymes were studied in the brain cortex after short (15 min) cerebral ischemia and reperfusion (10 min) in rats. Conjugated dienes (CD) and thiobarbituric acid-reactive substances (TBARS) were significantly elevated in the group of rats with ischemia followed by reperfusion in comparison to the ischemic animals. Superoxide dismutase (SOD) activity significantly increased in the group of animals with ischemia and reperfusion. No significant changes in the activities of glutathione peroxidase (GP) were observed. Stobadine administered before ischemia or before reperfusion decreased the level of TBARS. Stobadine probably prevents malondialdehyde (MDA) formation from hydroperoxide or might elevate the activity of aldehyde dehydrogenase. In contradiction to the findings after long-lasting (4 h) ischemia and subsequent reperfusion, no decrease in the concentration of CD or in the activity of SOD or GP was found.


Phase response curve to anisomycin in tau mutant hamsters. Administration of the protein synthesis inhibitor, anisomycin, to wild type hamsters produces phase shifts in their circadian rhythms that have similarities to shifts produced by non-photic behavioral stimulation. A mutation that shortens the period of rhythms in hamsters results in altered responsiveness to non-photic input. However, responses of the mutants to anisomycin are unaffected: their phase response curve (PRC) for anisomycin is similar to that of wild types. This suggests that 1) anisomycin is not acting on mechanisms specifically involved in non-photic behavioral phase shifting, and 2) the mutation affects the non-photic input pathway or the pacemaker itself at a point that is upstream from anisomycin's site of action.


Anti-muscarinic activity of a family of C11N5 compounds isolated from Agelas sponges. In a search for potential target sites for C11N5 compounds obtained from marine sponges of the genus Agelas we evaluated their interaction with muscarinic acetylcholine receptors from rat brain membranes. In competition experiments with 3H-QNB these compounds displayed the following rank order of potency: sceptrin greater than oroidin greater than or equal to dibromosceptrin greater than or equal to clathrodin. Sceptrin (50 microM) was shown to be a competitive inhibitor of 3H-QNB binding as revealed by Scatchard analysis. The results demonstrate the ability of these compounds to interact with multiple target molecules in the micromolar range.


Isometachromin, a new cytotoxic sesquiterpenoid from a deep water sponge of the family Spongiidae. Isometachromin (1), a new sesquiterpene-quinone that is related structurally to metachromin C (2), and the known compounds ilimaquinone (3) and 5-epi-ilimaquinone (4), were isolated from a deep water sponge in the family Spongiidae; the structure of isometachromin was elucidated by spectral methods. Isometachromin exhibits in vitro cytotoxicity against the human lung cancer cell line A549 (IC50 = 2.6 micrograms/ml), but not against P388 murine leukemia (IC 50 > or equal to 10 micrograms/ml) and also exhibits antimicrobial activity.


Differential olfactory perception of enantiomeric compounds by blind subterranean mole rats (Spalax ehrenbergi). The behavior of mole rats (Spalax ehrenbergi) near pairs of enantiomeric compounds was examined in 901 two-choice experimental tests. Positioning of the nest and food store and the preferred location of the tested animal were used to assess attraction or aversion to the tested odorants. The results indicated that mole rats respond differentially to odors of stereoisomers (enantiomers of carvone, citronellol, and fechone). They responded to one enantiomer of each tested pair but were indifferent to or did not smell the other. Both sexes were attracted to the odor of R-(-)-carvone and repelled by the odor of (+)-citronellol. Females were attracted to the odor of (-)-fenchone while males had no preference. By contrast, all animals were indifferent to or did not smell the odor of S-(+)-carvone, (-)-citronellol, and (+)-fenchone. Further research to distinguish between these alternatives (indifference vs hyposmia/anosmia) is suggested.


Adhesive grass spikelet with mammalian hair in Dominican amber: first fossil evidence of epizoochory. Discovery of a female spikelet of the grass genus Pharus (Gramineae: Bambusoideae: Phareae) in association with mammalian hair in Dominican Republic amber provides the first fossil evidence of epizoochory. Hooked macrohairs on the lemma of the spikelet show that morphological modifications in grasses for dispersal by attachment to the surface of animals were present in the Late Eocene. The fossil also represents 1) the second-oldest undoubted macrofossil record of the Gramineae, 2) the earliest record of a fossil grass that can be assigned to an extant genus, 3) the earliest undoubted record of a member of the bamboo subfamily and 4) the only known fossil of Pharus.


Ancylostoma ceylanicum infection in female hamsters: an observation on altered reproductive function. Reproductive performances of female hamsters were investigated during Ancylostoma ceylanicum (hookworm) infection. Animals having the highest levels of infection (34.96 +/- 1.11 worms) showed degenerative changes in the reproductive system. Ovaries of infected animals contained a few primary or secondary follicles. On cocaging with males of proven fertility, only 7-8% (80% in controls) of the infected females mated but did not conceive as evidenced by the absence of corpora lutea or implantation sites on day 10 postcoitum. Animals with low worm burdens (5.94 +/- 0.65 worms), however, showed almost normal fertility. The uterine weight bioassay and compensatory ovarian hypertrophy suggest strong suppression of pituitary gonadotrophin contents in infected females. Resorptive effects on the pregnancy outcome of infected female hamsters were also recorded.


Lipid-laden white blood cells in the circulation of patients with coronary heart disease. The total lipid content of white blood cells from healthy donors and patients with angiographically documented atherosclerosis of coronary arteries (CHD patients) was determined by flow cytofluorometry. The cells of donors were homogeneous with respect to intracellular lipid level. However, two subpopulations of white blood cells were identified in CHD patients based on their lipid content. The first population was identical to cells of donors with regard to lipid content, whereas cells of the second subpopulation (10 to 60% of the total cell number) had a four- to eightfold greater amount of intracellular lipid. The main classes of lipids accumulated in these cells were triglycerides and cholesteryl esters. It is postulated that the occurrence of lipid-laden cells in the blood may be used as an indicator of the presence of atherosclerotic lesions in human coronary arteries.


Tight junction permeability and liver plasma membrane fluidity in lithocholate-induced cholestasis. The present study correlated the reversibility of bile flow (BF) impairment with biochemical and morphological changes in the liver after injection of a cholestatic dose (12 mumole/100 g body weight) of lithocholic acid (LCA). BF declined maximally at 60 min but recovered totally at 210 min after LCA treatment. During the cholestatic period, there was an increase in tight junction permeability as measured by the bile to plasma (B/P) ratio of inulin and using lanthanum as a tracer. Cholesterol content and the cholesterol/phospholipid ratio in liver plasma membranes (LPM) were augmented while the fluidity of bile canalicular membranes (BCM) was decreased at 30 and 60 min after LCA injection. These changes in cholesterol content and membrane fluidity seemed to be correlated with LCA incorporation in LPM; their reversal at 120 min preceded the recovery of BF (210 min). Some biochemical disorders were evident after LCA injection, but they did not correlate with the variation in BF. These data suggest that increased tight junction permeability and decreased BCM fluidity are important pathogenic steps in LCA-induced cholestasis.


Antiatherosclerotic effects of antioxidants are lesion-specific when evaluated in hypercholesterolemic New Zealand white rabbits. Oxidative modification of LDL may represent an initiating event in the formation of monocyte-macrophage foam cells, a major cell present in fatty streaks and atherosclerotic fibrous plaques. Therefore, we studied the effect of such antioxidants as probucol (500 mg/kg) and vitamins E and C (500 mg/kg each) on the regression of induced iliac-femoral lesions and progression of naturally occurring thoracic aortic fatty streak lesions in hypercholesterolemic New Zealand White rabbits. Following an initial 9-week lesion induction phase, both therapies were evaluated for 8 weeks. Probucol lowered plasma cholesterol 47% while vitamins E and C had no effect on plasma cholesterol. Probucol decreased the cholesteryl ester (CE) content of the thoracic aorta by 31% without changing the thoracic aortic lesion coverage. Vitamins E and C decreased thoracic aortic CE content by 40% and lesion coverage by 46%. Neither probucol nor vitamins E and C altered the CE content, lesion size, or macrophage/lesion ratio of the iliac-femoral artery. Thus, we conclude that the effects of antioxidants are specific to the stage of atherosclerotic lesion development. Antioxidant therapy alters the progression and cholesteryl ester enrichment of diet-induced thoracic aortic fatty streaks but has no effect on the progression and/or regression of more complicated injury-induced iliac-femoral lesions.


Detection of migration stimulating activity in wound fluid. Fetal skin fibroblasts produce a soluble "migration stimulating factor" (MSF) which is not made by their normal adult counterparts. MSF stimulates the migration of adult skin fibroblasts into 3D collagen gels, thus providing the basis of a convenient bioassay for its presence. We have previously reported that MSF stimulates hyaluronic acid (HA) synthesis by adult skin fibroblasts and that this effect on matrix deposition appears to be responsible for the observed increase in cell motility. In the present study, wound fluid samples were collected from 18 patients undergoing surgery for various nonmalignant conditions and these were then fractionated according to the protocol used to isolate MSF from fetal fibroblast-conditioned medium. Detectable levels of migration stimulating activity were present in 17/18 (94%) of these samples. Paired serum samples obtained both pre- and postoperatively from five patients with positive wound fluid samples were also analyzed for MSF activity; such activity was found in only 1/5 (20%) of the preoperative and 0/5 (0%) of the postoperative serum samples. These data suggest that the MSF present in wound fluid is not derived from a plasma transudate or from platelet degranulation, but may reflect the transient and localized reinitiation of MSF production by adult fibroblasts in response to wounding. Taken together with previous observations regarding the effect of MSF on fibroblast migration and HA synthesis, our data suggest a possible physiological function of MSF in the wound healing response. Previous studies have revealed that MSF is produced by a subpopulation of apparently persistent fetal-like skin fibroblasts obtained from breast cancer patients and is also found in the serum of these individuals. Wound fluid and serum samples were accordingly collected from patients undergoing surgery for various types of malignant conditions or with a previous history of cancer; detectable levels of MSF activity were found in 8/10 (80%) of these wound fluid samples, 2/3 (66.6%) of the preoperative, and 3/3 (100%) of the postoperative paired serum samples. These findings suggest that the presence of detectable serum levels of MSF is not restricted to breast cancer and may be a general feature of malignant disease.


The ecology of Cryptococcus neoformans.

The ecology of Cryptococcus neoformans. Environmental isolations have established that Cryptococcus neoformans var. gattii serotype B appears to have a specific ecological association with Eucalyptus camaldulensis. The global distribution of the tree appears to correspond to the epidemiologic distribution of cryptococcosis caused by C. neoformans var. gattii. The epidemiology of cryptococcosis can primarily be explained by exposure to an infective aerosolized inoculum, such as basidiospores released from specific host plants and/or desiccated blastoconidia (yeast cells) disseminated from accumulations of dried pigeon dung. The ecology of C. neoformans still remains largely unresolved, studies on the host-parasite interaction between serotype B and E. camaldulensis are still in progress, and extensive environmental searches are now underway to determine the natural habitats of serotypes A, C and D.


Incidence of dermatophytoses in rabbit farms in Catalonia, Spain, and its repercussion on human health. Over the past decades there has been an important increase in the incidence of dermatophytoses in humans as a result of contact with animals, although etiological agents can vary as can the animals transmitting the disease. A large-scale study was carried out in 220 farms raising rabbits for consumption. Most of the farms (85%) were located in the autonomous community of Catalonia (Spain). Mycological studies showed that 79.5% of the rabbits were infected with Trichophyton mentagrophytes var. granulosum. Microsporum canis was isolated in only two animals, which had been imported from France. Healthy animal carriers were detected in 3.2% of the apparently non-infected farms. T. mentagrophytes were also found in samples taken from rabbits' nests and from the surrounding environment of the two infected farms. In a survey carried out among the staff responsible for the care of the animals, 77% of those working on infected farms suffered or had suffered dermatophytic lesions. This was confirmed in 8 out of 10 cases sampled. Attention is drawn to the high incidence of dermatophytoses in rabbits on farms and the importance of T. mentagrophytes as the etiological agent of tinea in people in close contact with infected animals.


Morphological and biochemical variability of Microsporum canis strains. Seventy-two strains of Microsporum canis, of different origins, were examined from a morphological point of view and tested in relation to their hydrolytic activity on tyrosine, xanthine, casein, gelatin, their ureasic activity and their capacity to assimilate different nitrogenous substances. The morphological aspects, that vary within the M. canis isolates, were constant in the strains isolated from rabbits. A strain with particular features was isolated many times from the dogs and cats coming from the same breeder. In one case of pseudomycetoma, different isolates suggested the co-existence in animals of two different strains, one present on fur, the other responsible for deep lesions.


Genetic susceptibility to dermatophytosis. Clustering of cases of dermatophytosis suggests that inherited susceptibility may play a part in determining the epidemiology of some forms of this infection, notably tinea imbricata. Some studies of T. concentricum infection show that autosomal recessive susceptibility may provide an answer although this is not the case in all endemic areas. Further support comes from the association between dermatophytosis in man and inherited conditions such as atopy, chronic mucocutaneous candidosis and tylosis as well as experimental data showing that susceptibility to dermatophytosis in mice varies in different inbred strains. Possible mechanisms are discussed.


Inhibition of adherence of Candida albicans to acrylic by a chitin derivative. The purpose of this study was to assess the effect of a chitin derivative (CSE) on the adherence of Candida albicans to acrylic. Fungal adherence to acrylic dentures is considered an essential step in the development of denture stomatitis. Adherence of C. albicans to acrylic pieces (5 x 5mm) was assessed microscopically using a calibrated ocular objective and expressed as number of adherent yeasts/mm2 of acrylic. CSE was prepared from commercial chitin (crab shell) and from chitin isolated from C. albicans blastospores. The effect of both CSE types on the adherence of C. albicans to acrylic was examined in two experimental systems: CSE present during the adherence assay and acrylic pieces pretreated with CSE prior to the assay. Both CSE types exerted a significant inhibitory effect when tested in the two experimental systems. These findings are significant for possible prevention of denture stomatitis.


Immunohistologic diagnosis of systemic mycoses: an update. Fluorescent antibody, immunoperoxidase and gold-silver staining methods for the rapid and accurate diagnosis of systemic mycotic infections are currently performed in a few specialized laboratories. These methods have proved applicable to formalin-fixed, paraffin-embedded tissues, and are reliable for identifying therein antigens of infectious dimorphic, monomorphic filamentous, and yeast-like fungal pathogens, i.e., Aspergillus spp., Blastomyces dermatitidis, Candida spp., Coccidioides immitis, Cryptococcus neoformans, Fusarium spp., Histoplasma capsulatum, Paracoccidioides brasiliensis, Pseudallescheria boydii, and Sporothrix schenckii. Most of the available reagents are derived from multiple adsorbed polyclonal antisera. However, problems occur in the production of uniform and standardized species- or genus- specific antibodies. Monoclonal antibodies, although promising, have to date not eliminated these problems. Immunohistologic methods will become more routinely used in clinical laboratories as these problems are resolved and more sensitive and specific reagents become commercially available.


Phaeohyphomycosis caused by Bipolaris spicifera: an informative case. A case of phaeohyphomycosis caused by Bipolaris spicifera involving the brain and sinuses is presented. The patient survived following surgery and ketoconazole therapy, which successfully treated both the sinus and the brain infections.


Cerebral phaeohyphomycosis caused by Xylohypha bantiana, with a review of the literature. A 76-year-old male with chief complaints of back and right leg sciatica was hospitalized. His abdominal CT scan revealed lumber spondylitic stenosis. A laminectomy was performed. Postoperatively, he became febrile, aphasic and had grand mal seizure. A left craniotomy of the front abscess, seen in the CT scan, was performed. H and E stained smears of drainage revealed dematiaceous, septate hyphae. Cultures of the abscess drainage grew an olivaceous-grey fungus. Based on macro- and micro-morphological characters, growth at 42 degrees C, and exoantigenic analysis, the patient's fungus was identified as Xylohypha bantiana. Treatment with amphotericin B and 5-fluorocytosine was initiated. Despite surgical procedures and antifungal therapy, the patient's condition deteriorated and he died a few weeks later due to cerebral edema. The case reported here is the first microscopically, culturally, histopathologically and exoantigenically proven case of phaeohyphomycosis caused by X. bantiana in the province of Alberta and from Canada. A review of the literature on cases of X. bantiana infections has also been presented.


Entomophthoromycosis due to Conidiobolus. Entomophthoromycosis due to Conidiobolus coronatus is a granulomatous infection characterized by lesions that originate in the inferior turbinate, spread through ostia and foramina to involve the facial and subcutaneous tissues and paranasal sinuses. The majority of the cases have been described from areas of tropical rainforest in West Africa, agricultural and outdoor workers (aged 20-60 years) being the ones most frequently affected. The fungus is common in soil and decaying vegetation. Infection probably occurs by implantation of the spores of the fungus in nasal mucosa. C. incongruus is a rare agent of the disease, so far known only from two cases with lesions involving the pericardium, mediastinum, lungs, liver, oesophagus and jejunum. C. coronatus is known to cause a clinically similar disease in horses, mules, a dolphin and a chimpanzee. A characteristic histological feature is the presence of thin-walled, broad, often septate hyphae or hyphal fragments with a thick eosinophilic sheath, frequently phagocytosed within giant cells. The fungus is known to produce in vitro several enzymes, e.g., elastase, esterase, collagenase and lipase, which have a possible role in pathogenicity. A concentrated brain heart infusion culture filtrate antigen is useful for immunodiagnosis. Several drugs e.g., potassium iodide, cotrimoxazole, amphotericin B, ketoconazole and itraconazole have been tried with varying success. Investigations on the immunology of disease and the role of proteases and lipases in the pathogenesis of infection is an important area of further research.


Protothecosis: a report of two cases in Japan and a review of the literature. Protothecosis is an emerging opportunistic infection caused by species belonging to the genus Prototheca. Two Japanese cases of protothecosis are documented with a critical review of the literature. A current perspective concerning the microbiology and disease entity of protothecosis is described in detail.


In vitro sensitivity of Penicillium marneffei and Pythium insidiosum to various antifungal agents. Ten isolates of Penicillium marneffei and eight of Pythium insidiosum were tested for their in vitro sensitivity to amphotericin B, hamycin (a polyene heptaene), two water-soluble analogs of amphotericin B and hamycin, namely, JAI-Amb, and JAI-hamycin,5-fluorocytosine, fluconazole, itraconazole,ketoconazole and miconazole. Itraconazole manifested the strongest activity against all of the 10 isolates of P. marneffei and would be the drug of choice in the treatment of penicilliosis due to P. marneffei. The polyene antibiotics amphotericin B and hamycin and their water-soluble analogs showed no appreciable activity against P. insidiosum. Pytium insidiosum isolates were sensitive to fluconazole, ketoconazole, and miconazole. Miconazole exhibited the strongest in vitro activity against all of the 8 isolates of P. insidiosum, followed by ketoconazole.


Drug resistance in human pathogenic fungi. Since the therapy of the mycoses, particularly the systemic mycoses, is relatively long-term in nature, emergence of resistance to antifungal drugs during the treatment of period would be of considerable clinical importance. However, most reports of resistance to antifungal agents among human pathogenic fungi indicate that naturally-occurring resistance is very rare, and that the induction of resistant mutants or variants is much more difficult to achieve in vitro and in vivo than with bacteria. As a matter of fact, amphotericin B and some other classic antifungals have not as yet posed a broadly significant problem relative to drug resistance despite their widespread and frequent use. Fungal resistance has thus received little attention, in contrast to the critical importance of bacterial resistance frequently caused by a variety of antibacterial chemotherapeutic agents, until a single exception to this generalization arose with the advent of flucytosine. This new development has aroused great interest in the problem of fungal resistance among the scientists involved with medical mycology. It is generally believed that fungi, like bacteria, are intrinsically capable of developing resistance to antifungal agents. As illustrated by flucytosine, inherently resistant mutants to antifungals occur within sensitive strains of human pathogenic fungi with significant frequency. Given the relatively high degree of such primary resistance, these mutants should develop secondary resistance during therapy, thus resulting in considerable limitations in the clinical usefulness of the antifungals. Virtually, all unsuccessful cases of mycoses treated with some of the recently exploited antifungal drugs, albeit scarce to date, would obviously be attributable to the occurrence of secondary resistance. The exploitation of new antifungal drugs thus requires investigations of their resistance as one of the most important research projects to be undertaken before receiving approval for use on humans. This paper reviews from various aspects the literature on resistance to various classic and novel antifungal agents among human pathogenic fungi. The resistance of some nonpathogenic fungi to these agents will also be described from genetic and biochemical points of view.


Surveillance and treatment of liver transplant recipients for candidiasis and aspergillosis. Between June 1988 and May 1991 88 orthotopic liver transplants and 1 liver and pancreas transplant were performed at the Liver Transplantation Department of the Ospedale Maggiore of Milan. All the patients underwent mycological surveillance and received antifungal prophylaxis with oral amphotericin B (6000 mg/day) or oral or intravenous fluconazole (200 mg/day) from the time of their transplant. The incidence of Candida colonization was 67%. Fluconazole was superior to oral amphotericin B in the treatment of C. albicans colonization (9/9 vs 6/15), but less effective in the treatment of colonization by other Candida spp. (0/3 vs 3/3). Deep-seated candidiasis developed in 5 patients, caused by C. albicans in 4 cases and C. krusei in 1. C. albicans infection resolved rapidly with fluconazole in 2 subjects, with intravenous amphotericin B alone in 1, and with amphotericin B plus flucytosine in the other. On the contrary, C. krusei infection did not respond to treatment with amphotericin B combined with flucytosine. Aspergillosis was diagnosed in 11 patients, of whom 4 died from invasive aspergillosis, despite 15 and 26 days of amphotericin B treatment in 2. In another patient invasive aspergillosis, diagnosed a few hours before retransplantation, improved with liposomal amphotericin B, but this man died from cytomegalovirus infection one month later. Aspergillosis was eradicated by itraconazole in 4 other patients and by topical amphotericin B in 2 whose infection was localized to surgical wound.


Distribution and antimicrobial susceptibility of Rhodococcus equi from clinical specimens. Rhodococcus equi, an unusual gram positive aerobic actinomycete, was first described as a respiratory pathogen of livestock in 1923. Reports of human clinical illness have emphasized R. equi as a cause of invasive pulmonary infection in severely immunocompromised patients and, recently, have implicated it as a cause of pneumonia, bacteremia and disseminated infection in HIV-infected patients. To determine the distribution of R. equi we evaluated 107 isolates referred to the Centers for Disease Control (CDC) during the period January 1973 through December 1990. The sites of these 107 isolates (101 patient and 6 animal isolates) were: blood (32 isolates), sputum (30), lung tissue (13) and other site (32). Before 1983, when the first R. equi isolate from an HIV-infected patient was received, CDC received a total of 52 patient isolates. In addition, during this 10 year period, R. equi isolates were received from more than one site from only one patient. However, during the two year period 1989-1990, we identified 8 patients with underlying HIV infection and R. equi pneumonia who accounted for 29 of 35 (83%) R. equi patient isolates; 6 of these patients also had bacteremia and three died with disseminated R. equi infection. No isolates were resistant to amoxicillin-clavulanate, ampicillin-sulbactam, gentamicin or imipenem, and few (less than 5%) isolates were resistant to erythromycin, rifampin, tetracycline, and trimethoprim-sulfamethoxazole. These results suggest that HIV-infected patients, in particular, are predisposed to develop invasive pulmonary, fatal disseminated R. equi infection (or both), and appropriate antimicrobial susceptibility testing of clinical isolates may improve the effectiveness of therapy of R. equi-infected patients.


DNA relatedness, karyotyping and gene probing of Candida tropicalis, Candida albicans and its synonyms Candida stellatoidea and Candida claussenii. Isolates of Candida albicans with varied phenotypes, including sucrose-negative variants (C. stellatoidea, serotypes A and B) and avirulent germ tube-negative forms (C. claussenii) showed significant (greater than 90%) DNA relatedness to classical C. albicans, but insignificant relatedness to C. tropicalis and sucrose-negative C. tropicalis. A transverse alternating-field gel electrophoresis procedure (TAFE) showed discrete karyotype patterns among the phenotypic variants of C. albicans including the sucrose-negative C. stellatoidea. The number of chromosome-sized DNA bands for C. tropicalis (7 bands) were within the range of bands observed for C. albicans (5 to 10 bands). The general DNA-migration pattern for C. albicans appeared distinct from that of C. tropicalis. An aspartyl proteinase (PrA) gene probe from C. albicans hybridized with chromosomal DNA from C. albicans, C. claussenii and C. stellatoidea but not with that from C. tropicalis.


Is Pneumocystis carinii a deep mycosis-like agent? Pneumocystis carinii is a widespread eukaryotic microorganism found in the lungs of healthy mammals, including humans. It is able to proliferate extensively in the alveoli, becoming an important agent of severe pneumonitis in immunosuppressed hosts, especially in persons suffering from AIDS. The taxonomic position of P. carinii is uncertain. Typical cytoplasmic organelles of eukaryotic cells have been found and described in the parasite. Biochemical research is hindered by the lack of an efficient in vitro culture system. Results of comparative study of nucleic acid sequences suggest that Pneumocystis is a fungus. However, ultrastructural, biochemical and nucleic acid homology insights appear as clearly insufficient to class Pneumocystis. Pneumocystis infection might be acquired, as deep mycoses, from environmental sources through the respiratory tract. Thus, the hypothesis of an environmental stage of the parasite must be considered. Pneumocystis might be seen as a widespread pathogenic dimorphous fungus. As fungal agents, P. carinii is able to disseminate from the infected lung to other organs. However, deep mycoses and pneumocystosis induce different histopathological changes in the host. Furthermore, deep fungal infections, unlike pneumocystosis, cannot be transmitted from one infested host to another one. Beside these two aspects, pneumocystosis shares many features with deep mycoses. Research on the epidemiology of pneumocystosis is needed.


Anaerobic yeast killer systems.

Anaerobic yeast killer systems. The influence of anaerobic conditions on the expression of the killer phenomenon of several yeast isolates belonging to recognized killer systems coded by different genetic determinants (Pichia spp., Kluyveromyces lactis, Saccharomyces cerevisiae) was studied. Anaerobiosis influenced the activity of killer toxins from some individual isolates of the genera Pichia and Saccharomyces on sensitive strains of P. anomala, K. lactis and Candida albicans. However, no influence was detectable on a S. cerevisiae sensitive isolate. Thus, anaerobic conditions seem to interfere more with the metabolic process of sensitive strains than with toxin production by killer yeasts. The selection of a panel of killer yeasts, able to display their activity against reference sensitive yeast isolates under anaerobic conditions in a medium that favored the growth of anaerobes, allowed the use of the killer system to type Bacteroides fragilis isolates for epidemiological purposes.


Five years of sentinel surveillance of acute respiratory infections (1985-1990): the benefits of an influenza early warning system. For the last five years, the Brussels Institute of Hygiene and Epidemiology has been involved in the surveillance of acute respiratory infections (ARI). The four indicators used (number of encounters of ARI by GP's/100 encounters, virus isolations, absenteeism and mortality) are discussed. A regression procedure is applied to the data collected by a sentinel network of general practitioners (GP's). This procedure permits the baseline to be visualized and an epidemic threshold to be determined in order to recognize early an influenza outbreak. The traditional use of flu-like illnesses as an indicator might be improved by the addition of non-specific ARI which are more precocious, especially in children. The criteria for an accurate definition of an influenza epidemic are discussed. The same mathematical model can be used for the analysis of mortality linked with an outbreak. It shows that the last epidemic in the winter 1989-1990 was responsible for about 4900 deaths directly or indirectly related to influenza.


Immunization of elderly volunteers with the 1988-89 inactivated whole influenza vaccine: assessment of antibody responses by haemagglutination inhibition and single radial haemolysis tests. The immunogenicity of inactivated whole trivalent influenza vaccines (A/Taiwan/1/86 (H1N1), A/Sichuan/2/87 (H3N2), and B ijing/1/87) recommended for the 1988-89 winter season was evaluated in 236 elderly (mean age 71 years) high risk volunteers. An overall significant increase in the number of subjects with protective haemagglutination inhibiting (HI) antibodies (titer > 1:40) against vaccine components was observed after vaccination. Nevertheless, a percentage of individuals (ranging from 56% to 62%) remained without protective antibodies and the number of people showing a positive response was limited (from 32% to 41%). By the comparative analysis of the results obtained examining the presence of protective levels of antibody in the sera from 91 volunteers using HI versus the single radial haemolysis (SRH) test, we obtained evidence for a higher sensitivity of SRH technique especially against B antigen.


The role of breast self-examination in early breast cancer detection (results of the 5-years USSR/WHO randomized study in Leningrad). A randomized population-based study has been carried out since 1985 in Leningrad in order to evaluate the efficacy of breast self-examination (BSE) in early breast cancer detection. The population under study covers 120,310 women aged 40-64 years with no history of breast cancer. About half of these women were exposed to BSE training (60,221) and 60,098 women constituted the control group. BSE teaching was carried out on a person-to-person basis and each patient received the BSE calendar. BSE education sessions resulted in a higher frequency of visits to specialists with complaints about "pathology" of the breast, a higher rate of referral to a specialized institution for an examination, and a higher number of excision biopsies due to a benign lesion (RR = 1.5; 95% C.I. = 1.1 - 1.9) as compared with the control group. As a result of examination, 190 breast cancer patients in the BSE group and 192 patients in the control group were detected. Comparisons of patients from both groups with regard to the size of primary tumor and the incidence of metastatic lesion in the regional lymph nodes showed no differences. The study is ongoing and all cases of breast cancer in the BSE group will be registered up to 1994 and followed-up to 1999; information will then be available on the impact of BSE upon breast cancer mortality.


Maternal diabetes: the risk for specific birth defects. We studied the risk for specific birth defects among infants of mothers with gestational and chronic diabetes using data collected by the Spanish Collaborative Study of Congenital Malformations (ECEMC). For the years 1976 to 1985, we identified 10,087 infants with malformations and 9,994 control infants; 155 of the case infants and 89 of the controls were born to diabetic mothers. The crude odds ratio for any minor or major defect and insulin-treated diabetes was 5.5 (95% CI = 1.2, 24.8), and for major malformations it was 8.7 (95% CI = 1.8, 34.7). The risk for defects involving the central nervous system (CNS), skeletal system and cardiovascular system were significantly increased. Infants of non-insulin-treated diabetic mothers were 2.9 times more likely to have a major congenital birth defect (95% CI = 1.2, 7.2). The crude odds ratio for any major or minor defect and mothers with gestational diabetes requiring insulin was 1.9 (95% CI = 1.1, 3.4). Similar risk was observed for major defects (OR = 1.9, 95% CI = 1.0, 3.7). These results suggest that infants of insulin-treated diabetic mothers have an increased risk of developing malformations of the CNS, cardiovascular system and skeletal system. We also found an increased risk for specific defect categories among infants of mothers with gestational diabetes treated with insulin.


Risk factors for atrial septal defect.

Risk factors for atrial septal defect. The possible effect of environmental factors during pregnancy on the occurrence of atrial septal defect (ASD-secundum) in the offspring was studied in 50 cases and 756 controls. The cases represented all verified ASDs in Finland during 1982-1983. The controls were randomly selected from all infants born during the same period. Case and control mothers were interviewed by midwives using a structured questionnaire approximately three months after delivery. Congenital heart disease was more prevalent among parents of cases than those of controls. Maternal alcohol consumption during the first trimester of pregnancy appeared to double the risk of atrial septal defect (OR = 1.9, CI98 = 1.1 - 3.4). Maternal exposure to chemicals at work during the first trimester was more prevalent among the ASD-group (40.0%) than the control group (26.2%). The risk of ASD was not associated with maternal smoking, or coffee, tea or acetosalicylic acid consumption. Maternal exposure to video display terminals, microwave ovens, organic solvents, anesthetic gases, pesticides or wood preservatives during the first trimester of pregnancy were not associated with the risk of an atrial septal defect. It is concluded that some common physical and chemical exposures during early pregnancy should not necessarily be considered risk factors for atrial septal defect.


Incidence of congenital toxoplasmosis in live Guatemalan newborns. Five hundred and fifty samples of blood collected from the umbilical cords of an equivalent number of newborns were analyzed for serological evidence of congenital toxoplasmosis based on the detection of IgG and IgM. Six newborns presented serological evidence of congenital toxoplasmosis (IgM > 1:5, < 80 IU/ml), which represents an incidence of 10.9 per 1000 live births. During pregnancy four of the mothers of these six newborns were asymptomatic, whereas the other two mothers presented non-specific signs and symptoms. The six newborns did not present positive signs of acute toxoplasmosis at birth. Three false positive were identified, all secondary to the presence of a rheumatoid (RF) and/or antinuclear factor (ANF). And in one of them the diagnosis of congenital syphilis was confirmed. The percentage of women that tested serum positive for IgG antibodies increased with age, with 55.8% of the pregnant women testing serum-negative, therefore carrying the risk of acquired toxoplasmosis in future pregnancies.