Long term follow-up of a case of Kasabach-Merritt syndrome successfully treated with radiotherapy and corticosteroids.

Kasabach-Merritt syndrome is the association of thrombocytopenia, spontaneous bleeding, and enlargement of a haemangioma. It is caused by an intense, self-perpetuating process of clot-formation and lysis within the abnormal vascular channels of the haemangioma, and results in consumption of platelets and clotting factors. Treatment involves ablation of the lesion with or without pharmacological manipulation of the coagulation and fibrinolytic systems. No single therapeutic modality is universally successful but a combination of radiotherapy and corticosteroids can result in a dramatic, immediate response with minimal long term complications.


Interaction between verbal and gestural language in progressive aphasia: a longitudinal case study.

The objective of this longitudinal study is to investigate the on-line interaction between praxis and linguistic abilities in a progressive aphasia case. During 3 years of evolution, procedural discourse of a progressive aphasic patient was videotaped five times, allowing us to analyze the progression of both language and gestural production as well as the interaction between these two. We anticipated that, in the absence of apraxia, the patient would compensate for her speech deficit by producing progressively more and more meaningful gestures. Our compensatory hypothesis was confirmed but the compensation was not as efficient as one would expect given the absence of apraxia. With the progression of the speech deficit, the patient could not replace some verbs by pantomimes that were otherwise accompanying her discourse in the preceding testing sessions. We suggest that such a compensatory ability may constitute one important characteristic of the progressive aphasia syndrome.


Impairment of temporal organization of speech in basal ganglia diseases.

Absolute and relative speech timing were examined in patients suffering from Parkinson's, Huntington's, and Wilson's disease. The task was to speak a standard sentence 10 times, first slowly, and then successively faster up to maximum rate. All patient groups had low maximal speech rates and showed decreased variability of speech rate. The duration of pauses between words was the same as in normals and the relative time structure of the test sentence was basically preserved. For comparison, two cases with nonfluent aphasia had even slower speech rates, large increases in pause duration, and major changes in relative speech timing. The results show the same type of alterations of the temporal organization of speech as those characteristic for rapid alternating limb movements in such patients. They support the view that the speech and skeletomotor systems share common neural control modes despite fundamental biomechanical differences. The common denominator between the speech and the skeletomotor disturbances in basal ganglia diseases may be the undamping and slowing of a fast central oscillator.


Dissociation of mechanisms of reading in Alzheimer's disease.

The role of spelling-to-sound correspondence rules in oral word reading was investigated by asking patients with Alzheimer's Disease (AD) and normal controls to read aloud pronounceable letter strings that do not happen to be real words. These pseudowords were of two types: those that have orthographically similar "neighbors," and those that have no neighbors. The patients with AD were mildly impaired relative to the normal controls in reading pseudowords with neighbors, but were markedly impaired in reading pseudowords with no neighbors. The results are interpreted as favoring a model of reading in which words and pseudowords are normally read via the same lexical mechanism. An ancillary route involving the conscious application of spelling-to-sound rules is available only to cognitively intact readers.


Dissociation between linguistic and nonlinguistic gestural systems: a case for compositionality.

This paper addresses the issue of the separability of disorders of sign language from disorders of gesture and pantomime. The study of a left-lesioned deaf signer presents one of the most striking examples to date of the cleavage between linguistic signs and manual pantomime. The left-hemisphere lesion produced a marked sign language aphasia disrupting both the production and the comprehension of sign language. However, in sharp contrast to the breakdown of sign language, the ability to communicate in nonlinguistic gesture was remarkably spared. This case has important implications for our understanding of the neural mediation of language and gesture. We argue that the differences observed in the fractionation of linguistic versus nonlinguistic gesture reflect differing degrees of compositionality of systems underlying language and gesture. The compositionality hypothesis receives support for the existence of phonemic paraphasias in sign language production, illustrating structural dissolution which is absent in the production of pantomimic gesture. Understanding the neural encoding of compositional motoric systems may lead to a principled anatomical account of the neural separability of language and gesture. This case provides a powerful indication of the left hemisphere's specialization for language-specific functions.


Hemispheric dissociation in judging semantic relations: complementarity for close and distant associates.

In two lateralized tachistoscopic experiments, we presented (i) pairs of nouns with close or distant semantic associations or (ii) pairs of nouns which were randomly matched and later rated by the subjects as to their semantic distance. In both experiments, words presented to the right visual field were more frequently judged as semantically close in meaning than words presented to the left visual field (LVF), whereas words presented to the LVF were more frequently judged as semantically distant. The results are discussed in relation to hemispheric language functions and current models of cerebral laterality.


Educational level and the word frequency effect: a lexical decision investigation.

It is known that speed and accuracy in recognizing words are constrained by the frequency of occurrence of these words ("frequency effect"). This study examines the relationship between educational level and the word frequency effect. We postulated that individual exposure to words that are rated lower in frequency tables should be greater among subjects with higher education and therefore hypothesized that the magnitude of the frequency effect should not be as marked within such a population as among subjects with a lesser educational level. A total of 40 neurologically healthy adults, half with an average of 18 years of formal education and the other half with an average of 11 years, participated in a lexical decision experiment. Results confirmed our hypothesis; that is, significant frequency effects on reaction times were obtained in both groups but this effect was of greater magnitude for the less educated as opposed to the more educated subgroup. The significance of this finding is discussed by reference to current models of word recognition.


Phrenological studies of aphasia before Broca: Broca's aphasia or Gall's aphasia?

The history of aphasia is usually taken to begin with Broca's (1861a,b) discovery of the correlation of aphemia with damage to the posterior inferior portion of the frontal lobes and the subsequent relation to left hemisphere. That there were prior, even biblical, references to aphasia is not in dispute, nor that, according to Benton and Joynt (1960), almost all the clinical forms of aphasia had been described prior to 1800. The significance of Broca's case studies, therefore, lies in the association of motor aphasia with focal pathology in the frontal lobe. This paper examines the status of aphasia localization prior to Broca, and, specifically, the extent to which Broca's discovery may have been anticipated by the phrenologists.


Self-repair in dialogues of schizophrenics: effects of hallucinations and negative symptoms.

This paper concerns the discourse features of verbal hallucinations and negative symptoms of schizophrenia. A total of 46 schizophrenics, varying in verbal hallucination and in negative symptoms status, and 22 controls were tested on the Reporter Test. The frequency with which they issued inadequate instructions, attempted to repair the inadequacies, and the success of repairs were compared. We observed that schizophrenics, on the whole, issued more wrong and incomplete instructions. This was in part related to their worse working memory, but it was not affected by verbal hallucinations or negative symptoms. We observed, further, that schizophrenics had no particular problems monitoring messages for inadequacies. We did find, however, that schizophrenics with verbal hallucinations had a specific problem with self-repairing wrong instructions. We interpret these results in the framework of Hoffman's (1986b) plan disruption-based model of verbal hallucinations; the Frith (1987) and Frith and Done (1988) internal monitoring model of positive and negative symptoms; and finally Frith and Frith's (1990) model of negative and positive schizophrenia and we use the results to specify the models.


Language lateralization in bilinguals: more not less is needed: a reply to Paradis (1990).

Paradis (1990) took an extremely pessimistic view of research to date looking at the lateralization of cerebral function in bilingual subjects. We, however, argue that some of his conclusions are, in fact, incorrect, while others speak more to the youth of the field. The problems in the latter body of work are now acknowledged and are being overcome. That the shortcomings are recognized seems to be more a case for satisfaction than of despair.


The Loch Ness Monster approach to bilingual language lateralization: a response to Berquier and Ashton.

To continue to search for a difference in language lateralization between bilinguals and unilinguals is at best a futile task in the face of all the clinical evidence to the contrary and inconsistent experimental findings. No amount of refining of dichotic, tachistoscopic, or concurrent task procedures, or statistical sophistication, is likely to tap the differential reliance on pragmatic cues that might very well exist in some nonbalanced bilinguals.


Polyclonal antibody localizes glia maturation factor beta-like immunoreactivity in neurons and glia.

A rabbit polyclonal antibody (91-01) was raised against recombinant human glia maturation factor beta (r-hGMF-beta). The antibody did not cross-react with a number of other growth factors on ELISA test. When compared with the monoclonal antibody G2-09 previously obtained, 91-01 immunoblotted the same protein band in rat brain extract. However, unlike G2-09 which immunostained only astrocytes and Bergmann glia, 91-01 stained neurons as well. Many but not all neurons in the central and peripheral nervous system were positive for GMF-beta. The larger cell population stained by the polyclonal antibody was most likely due to its increased sensitivity, although other explanations are possible. The presence of GMF-beta-like immunoreactivity in both neurons and glia raises the possibility of a wider range of cell-cell interaction than was previously considered.


Amygdala stimulation produces two types of hippocampal afterdischarges in the urethane-anesthetized rat.

How seizures spread during epileptiform events has been the subject of intensive investigation over the years. The present study explored the relationship between the amygdala and the hippocampus during afterdischarges. Stimulating electrodes were placed in the amygdala and CA3 regions on the left side in urethane-anesthetized rats. Recording microelectrodes were placed in the dentate gyrus on the left and the CA1 cell layer on the right. Afterdischarges were elicited by stimulus trains between 10 and 50 Hz to the amygdala. Most of the afterdischarges consisted of broad positive potentials in dentate gyrus and no shift of the DC potential. When the stimulus trains were repeated, the afterdischarge evolved, first by spreading to the contralateral side and then by the appearance of maximal dentate activation. The onset of maximal dentate activation was indicated by the appearance of bursts of large amplitude population spikes and a negative shift of the DC potential. These data demonstrate that two types of afterdischarges can be produced in the hippocampus of the anesthetized rat after amygdala stimulation. The observations support the hypothesis that maximal dentate activation represents synchronized reverberatory activity throughout the hippocampal-parahippocampal circuit and indicate that amygdala stimulation can access this circuit in the anesthetized animal.


Suppression of nociceptive responses in parafascicular neurons by stimulation of substantia nigra: an analysis of related inhibitory pathways.

A total of 166 neurons in parafascicular nucleus (PF) were studied, 85 from intact animals, 72 following dorsal spinal cord transection (D.Sp.C.X.), and 9 following complete transection of the spinal cord. Two patterns of nociceptive responses were identified following noxious stimulation and these responses were classified as 'nociceptive-on' and 'nociceptive-off' neurons, respectively. The effects of stimulating the substantia nigra (SNS) on the spontaneous and on the nociceptive evoked discharges were observed and compared in intact, D.Sp.C.X. and completely transected spinal cord rats. The results show that SNS significantly suppresses both the spontaneous and the nociceptive evoked discharges elicited by peroneal nerve stimulation. With an intact spinal cord, SNS suppressed both the spontaneous [-37 +/- 3.2% (P less than 0.05)] and the nociceptive evoked discharges [-52.8 +/- 2.8% (P less than 0.01)] of the 'nociceptive-on' cells respectively, while in the 'nociceptive-off' cells the same stimulation elicited an even more prominent suppression upon both discharges (-47.7 +/- 5.4%, P less than 0.01 and -64.9 +/- 5.0%, P less than 0.01), respectively. After D.Sp.C.X., the suppressive effects on the 'nociceptive-on' cells following SNS were diminished (-28.1 +/- 3.5% and -36.9 +/- 2.6%, respectively) but not abolished, while in the 'nociceptive-off' cells, the inhibitory effects on SNS were unchanged. In addition, the suppressive effects of SNS on the spontaneous activity of PF neurons in cases with completely cut spinal cords remains unchanged. These results suggest that SNS modulates the spontaneous and the noxious evoked responses of the PF neurons by way of supraspinal connections besides the previously described descending projecting pathways.


Effects of atropine on hippocampal theta cells and complex-spike cells.

The medial septal nuclei are essential for the naturally occurring hippocampal theta rhythm. Evidence that the rhythmic activity of the septum is carried via cholinergic afferents to the hippocampus has been: (a) the existence of a cholinergic septo-hippocampal projection, and (b) the sensitivity of one type of theta rhythm to antimuscarinic agents or cholinergic depletion. The muscarinic action of acetylcholine on pyramidal cells, however, is too slow to carry even a 4 Hz signal. Recent in vitro studies have confirmed a fast excitatory response by some hippocampal interneurons to muscarinic agonists. In urethane anesthetized rats, iontophoretic application of atropine to 17 hippocampal theta cells (presumed interneurons) during the theta rhythm, reduced their firing rates to an average of 24% of control rates. The effect of iontophoretic atropine application to 4 CA1 complex-spike cells (presumed pyramidal cells) was a selective elimination of their bursting activity with no significant effect on overall firing rate. The data suggest that: (1) interneuronal firing, during the hippocampal theta rhythm, is dominated by an excitatory cholinergic input and not by excitatory collaterals of pyramidal cells; and (2) somatic burst firing by CA1 pyramidal cells requires the presence of acetylcholine.


Expression of the non-proliferation-specific protein, statin, in grey matter neuroglia of the aging rat brain.

The monoclonal antibody, S-44, identifies statin, a 57 kDa nuclear protein which appears to be expressed exclusively in non-proliferating cells. We previously demonstrated that in the aging rat corpus callosum approximately one third of neuroglia are statin-negative, suggesting the existence of an unexpectedly large cycling glial compartment. In the present study, double-labeling of individual cultured astroglia with [3H]thymidine and the S-44 antibody provided direct evidence for the non-proliferative status of statin-positive cells. The S-44 antibody was used to immuno-localize statin and thereby determine growth fractions for neuroglia in various grey matter regions of 3-, 18-, and 33-month-old rats. The proportion of statin-negative (cycling) cells for the three ages combined ranged from about 24% in the molecular layer of the dentate gyrus to 38% in the molecular layer of the parietal cortex. In most regions surveyed total glial counts and proportions of statin-positive and -negative cells did not vary significantly as a function of advancing age. These results suggest that (i) as in corpus callosum, pools of cycling neuroglia in various grey matter regions are far in excess of those previously predicted by S-phase labeling with [3H]thymidine or BUdR, and (ii) ratios of proliferating-to-quiescent neuroglia are tightly regulated over much of the animal's adult life span. These conserved ratios may be used as markers of normal CNS senescence, and deviations thereof may indicate the presence and extent of intervening neuropathologic processes.


Effect of pentobarbital on cerebral regional venous O2 saturation heterogeneity.

Observed venous O2 saturation inhomogeneity in the brain implies a microregional imbalance in O2 supply/consumption. We hypothesized that this heterogeneity should be decreased by pentobarbital anesthesia through a reduction in regional metabolic heterogeneity. Male, Long-Evans, approximately 350 g rats were either anesthetized with 50 mg/kg pentobarbital (n = 10) or used as a conscious control group (n = 10, catheters inserted two hours earlier under ether anesthesia). In each rat, regional cerebral blood flow was determined by [14C]iodoantipyrine and regional arterial and venous O2 saturation were determined by microspectrophotometry. In the PB group, the mean blood pressure (107 +/- 7 Torr), heart rate (362 +/- 29/min), average cerebral blood flow (63 +/- 19 ml/min/100 g), and average cerebral O2 consumption (3.7 +/- 1.2 ml O2/min/100 g) were lower than those values in the conscious group (128 +/- 15, 474 +/- 44, 112 +/- 40, and 7 +/- 3), respectively. O2 extraction did not change after pentobarbital anesthesia. However, the dispersion of venous O2 saturation narrowed. The distribution of O2 saturations in 373 cerebral veins of anesthetized rats had a significantly reduced coefficient of variation [C.V. = 100 x (S.D./mean) = 13] as compared to a C.V. of 18 in 320 veins in conscious rats. Thus, pentobarbital anesthesia reduced the microregional venous O2 saturation inhomogeneity in the brain, creating a more uniform balance of oxygen supply and consumption.


Extracellular dopamine in the rat striatum during ischemia and reperfusion as measured by in vivo electrochemistry and in vivo microdialysis.

The effects of transient global forebrain ischemia and reperfusion on striatal extracellular dopamine levels were analyzed using both in vivo electrochemistry and in vivo microdialysis in urethane-anesthetized rats. Electrochemical records showed that extracellular dopamine levels increased once during the period of ischemia, and a second time during reperfusion. This biphasic pattern was not detected by microdialysis, probably because of the relatively low time resolution of this technique. Microdialysis provided evidence that the voltammetric signal was a measure of dopamine, and also allowed measurement of the metabolites dihydroxyphenylacetic acid and homovanillic acid, both of which decreased during ischemia. The biphasic dopamine pattern seen in rats is similar to that reported previously in gerbils, suggesting that it is a phenomenon common to transient ischemia and reperfusion across different species and models of transient global ischemia. This phenomenon may have important implications for therapeutic intervention in cerebral ischemia.


Role for protein synthesis in the neurotoxic effects of methamphetamine in mice and rats.

The mechanism by which the amphetamines damage selectively nigrostriatal dopaminergic neurons in experimental animals remains uncertain. The observation that neuronal cell death during embryogenesis involves an activation of gene expression and new protein synthesis, coupled with recent reports indicating that the amphetamines are capable of inducing neuropeptide biosynthesis, offers a possible clue as to their neurotoxic mechanism of action. Based on these considerations, we evaluated the effects of two different inhibitors of protein synthesis, cycloheximide and anisomycin, on the long-term, amine-depleting effects of methamphetamine (METH) in mice and rats. Both inhibitors were found to block the amine-depleting effects of METH in these species. In other experiments, cycloheximide did not affect the functional integrity of dopaminergic or glutamatergic neurons, transmitter systems previously implicated in the neurotoxic mechanism of action of METH. These findings raise the possibility that the neuronal-damaging effects of METH are mediated via a synthesis of 'neurotoxic' proteins.


Post-ischemic administration of bifemelane hydrochloride prohibits ischemia-induced depletion of the muscarinic M1-receptor and its mRNA in the gerbil hippocampus.

Parallel determinations of muscarinic cholinergic M1 receptor (M1-R) binding and of M1-R mRNA levels were carried out in the gerbil hippocampus 14 days after 5 min of transient ischemia. Both were reduced in the ischemic tissue to about 50% of the levels found in sham-operated controls, indicating that the late loss of M1-R is probably dependent on decreased synthesis. Three administrations of bifemelane hydrochloride (15 mg/kg, i.p., just after ischemia and 6 and 12 h later) completely prevented neuronal death in the hippocampus and ischemia-induced losses of hippocampal M1-R and its mRNA. Since vascular dementia may depend upon the ischemia-induced losses in cholinergic communication in the hippocampus, these findings suggest that it may be possible to prevent its occurrence by post-ischemic treatment with bifemelane hydrochloride.


Non-photic phase shifting of the circadian activity rhythm of Syrian hamsters: the relative potency of arousal and melatonin.

This study investigated the relative potency of melatonin and arousal as Zeitgebers in the non-photic phase shifting of circadian rhythmicity in the adult Syrian hamster. Animals held under dim red light (DD) exhibited robust free-running rhythms of wheel-running activity. Melatonin (1 mg/kg) or ethanolic saline vehicle, delivered manually by subcutaneous injection after removing the animal from its cage, resulted in phase advances of the activity rhythm. This effect was phase dependent, injections at CT 8 and 10 being effective (CT 12 = anticipated activity onset), whereas injection at CT 2, 6, 14 and 20 did not cause a shift. There was no significant difference between the magnitude or timing of phase shifts in response to injections of saline or melatonin. To determine whether the observed shifts were related to arousal of the animals induced by handling, a second group held under DD were fitted with chronic s.c. cannulae so that melatonin solution or vehicle could be delivered remotely at projected CT 10. Neither solution had any effect upon the free-running rhythm. However, when these animals received manual s.c. injection of saline or melatonin solution, they exhibited phase advances similar to those observed in Expt. 1. These results fail to support the hypothesis that melatonin can exert a chemically specific, acute phase-shifting action in the adult Syrian hamster. They do, however, demonstrate the potent effect of arousing stimuli upon the circadian clock in this species.


Ca(2+)-dependent release of [3H]GABA in cultured chick retina cells.

Depolarization by K+ (50 mM) of cultured chick retina cells released 1.14 +/- 0.28% of the accumulated [3H] gamma-aminobutyric acid (GABA) in the absence of Ca2+, but when 1.0 mM Ca2+ was present, the internal free calcium ion concentration [Ca2+]i rose by about 750 nM and the [3H]GABA release about doubled to a value of 2.22 +/- 0.2% of the total [3H]GABA. Nitrendipine (0.1 microM), a blocker of the L-type Ca2+ channels, blocked the [Ca2+]i response to K+ depolarization by about 65%, and the omega-Conotoxin GVIA (omega-CgTx) (0.5 microM), a blocker of the N-type of Ca2+ channels, inhibited by 27% the [Ca2+]i rise due to K+ depolarization. Parallel experiments showed that nitrendipine inhibits [3H]GABA release to the level observed in the absence of Ca2+, whereas omega-CgTx did not inhibit significantly the release of [3H]GABA. The results also show that the release of [3H]GABA due to K(+)-depolarization in the absence of Ca2+ can be totally blocked by 1-(2-(((Diphenylmethylene) amino)oxy)ethyl)-1,2,5,6-tetrahydro-3-pyridine-carboxylic acid hydrochloride (NNC-711), an inhibitor of the GABA carrier. However, in the presence of Ca2+, NNC-711 blocks the release only by about 66%, corresponding to the Ca(2+)-independent release. Thus, it is concluded that [3H]GABA is released in chick retina cells by the exocytotic mechanism, which is Ca(2+)-dependent, and by reversal of the carrier, which is Ca(2+)-independent, in much the same way as has been found for other GABAergic neurons.(ABSTRACT TRUNCATED AT 250 WORDS)


Topographical analysis of epileptiform potentials in rat somatosensory cortex: the interictal to ictal transition.

Large quantities of penicillin were applied to the face and forelimb region of rat somatosensory cortex, producing an epileptic focus with both electrographic and behavioral signs of seizures that regularly repeated over a period of several minutes. Epicortical potentials were recorded simultaneously from a 64 channel micro-electrode array (8 x 8 platinum electrodes) with inter-electrode distances of 0.5 mm, covering a 3.5 x 3.5 mm2 area centered on the penicillin injection site. Cluster analysis was used to classify successive epileptiform discharges into interictal, transitional, and ictal groups. Principal components analysis (PCA) was used to extract fundamental waveforms producing the spike complex in each group, and to estimate the locations and spatial extent of neuronal populations participating in epileptiform discharge. During all states of epileptic excitability, it was possible to account for over 90% of the variance in the epicortical potential waveforms using a model with only two spatially overlapping populations of cells. The location and spatial extent of the populations remained unchanged by the transition to seizures; the interictal and ictal states were distinguished only by changes in the timing and amplitude of potentials in the two putative neuronal populations. The present model, using only two stationary neuronal populations to reproduce all spatiotemporal patterns in the neocortical epileptogenic focus, is compared to models proposed by others in which epileptic discharge is thought to propagate sequentially through adjacent cortex. It is concluded that the initiation, maintenance, and termination of seizures in neocortex relies on mechanisms that are not necessarily reflected in changes in spatiotemporal interactions among epicortically recorded cell groups within the focus. These mechanisms may be distinguished from those responsible for the spread of seizures within neocortex.


Anatomical evidence for genetic differences in the innervation of the rat spinal cord by noradrenergic locus coeruleus neurons.

Pontospinal noradrenergic neurons located in the A5, A6 (locus coeruleus, LC), and A7 cell groups are the major source of the noradrenergic innervation of the spinal cord. We have recently examined the specific terminations of these three cell groups in the spinal cord and found that the LC provides the major noradrenergic innervation of the ventral horn, while the A7 and A5 cell groups innervate the dorsal horn and intermediate zone, respectively. However, the results of similar experiments from another laboratory have shown that noradrenergic neurons in the locus coeruleus primarily innervate the dorsal horn, while the A5 and A7 innervate the intermediate zone and the ventral horn. These conflicting results may be due to fundamental genetic differences between the rats used in our experiments (Sasco Sprague-Dawley) and those used by the other laboratory (Harlan Sprague-Dawley). This possibility was examined by determining the projections of coeruleospinal neurons in these two rat substrains using the anterograde tracer Phaseolus vulgaris leucoagglutinin. The results indicate that in Sasco rats the LC neurons project through the ipsilateral ventromedial funiculus and terminate almost exclusively in the medial part of laminae VII and VIII, the motoneuron pool of lamina IX, and lamina X. In contrast, LC neurons in Harlan rats project bilaterally through the superficial dorsal horn and the dorsolateral funiculus and terminate most heavily in dorsal horn laminae I-IV. In addition, the LC neurons of Sasco rats innervate cervical spinal cord segments more densely than lumbar spinal cord segments, while in Harlan rats the lumbar spinal cord is more densely innervated than the cervical spinal cord. These results indicate that the projections of coeruleospinal neurons in Sasco rats are fundamentally different from those in Harlan rats and suggest that noradrenergic LC neurons may have different physiological functions in these two rat substrains.


Muscimol infused into the medial septal area impairs long-term memory but not short-term memory in inhibitory avoidance, water maze place learning and rewarded alternation tasks.

These experiments investigated the effects of injections of muscimol (1 or 5 nmol), administered into the medial septal area prior to training, on memory tested at different retention delays after training in 3 tasks: an inhibitory avoidance task, a one-trial place learning task, and a rewarded alternation task. In all 3 tasks, intraseptal injections of muscimol did not impair memory performance at short retention delays, but impaired memory at the longer retention delays. These findings are consistent with the view that GABAergic regulation of the septohippocampal cholinergic system plays a selective role in the establishment of long-term memory.


gamma-Aminobutyric acid immunoreactive structures in the nucleus tractus solitarius: a light and electron microscopic study.

gamma-Aminobutyric acid immunoreactive perikarya and boutons in the nucleus tractus solitarius of the cat were examined at both the light and electron microscopic level. Immunoreactive neurones were found predominantly in the parvocellular subdivision of the nucleus tractus solitarius and to a lesser degree in all the other subdivisions of the nucleus tractus solitarius and the dorsal vagal motonucleus. All the immunoreactive perikarya observed were similar in size and morphology. gamma-Aminobutyric acid immunoreactive boutons were observed throughout the nucleus tractus solitarius. However, in contrast to its high content of immunoreactive perikarya the parvocellular subdivision contained the lowest density of immunoreactive boutons. Ultrastructural examination of immunoreactive boutons in the different regions of the nucleus tractus solitarius revealed that they formed synaptic specializations, predominantly with dendritic shafts, all of which were of the symmetric type. This pattern of innervation was observed throughout the medial, commissural, ventrolateral and parvocellular subdivisions of the nucleus tractus solitarius.


Medullary respiratory neurons in the guinea pig: localization and firing patterns.

The location and firing patterns of medullary respiratory neurons have been described in a small number of species. The cat has been the most widely studied species, but some potentially important differences have recently been noted in others. A more complete survey of species is required to determine the significance of these differences. We describe the location and firing patterns of respiratory neurons in the medulla of anesthetized, paralyzed and mechanically ventilated adult guinea pigs. Extracellular single-unit recordings were made from the medulla, their phase relationship with phrenic nerve activity used to define them as respiratory and their location marked with fast green. Respiratory units were concentrated ventrolateral to the nucleus tractus solitarius (NTS) and within and surrounding the nucleus ambiguus (NA), corresponding to the dorsal respiratory group (DRG) and ventral respiratory group (VRG) of the cat, respectively. Most DRG respiratory units were inspiratory, while the VRG contained equal numbers of inspiratory and expiratory units. The DRG and VRG both contained early, late and constant-frequency inspiratory and expiratory units. In general, these findings are similar to those in other mammalian species examined, consistent with these basic aspects of the respiratory network being highly conserved.


Characterization of interleukin-1 production by microglia in culture.

The production of interleukin-1 (IL-1) by cultured neonatal rat microglia was studied using the D10 cell assay. The results show that IL-1 was secreted in response to lipopolysaccharide (LPS) in a dose- and time-dependent fashion. IL-1 production was specific to microglia and was not induced in astrocytes. Indomethacin, which is known to modulate the release of IL-1 from monocytes, had no effect on LPS-stimulated microglia. Aging of the microglia from two weeks to 4 weeks in culture, however, reduced the release of IL-1 in response to LPS. Our data indicate that microglia are a major source of IL-1 and that the release of IL-1 depends on the presence of inflammatory mediators such as LPS and the age of the culture.


Distribution and relative density of p75 nerve growth factor receptors in the rat brain as a function of age and treatment with antibodies to nerve growth factor.

It is clear that nerve growth factor (NGF) has a role in the central nervous system. In order to begin to determine the possible roles of NGF in the CNS, neonatal rats were given daily subcutaneous injections of antibodies to NGF (ANTI-NGF) beginning at birth for a period of one month. By utilizing the monoclonal antibody, 192-IgG, which recognizes the p75 NGF receptor (NGFR), and standard immunohistochemical techniques we have localized p75 NGFR in variously aged ANTI-NGF-treated animals and compared the anatomic localization and relative density of the p75 NGFR immunoreactive (p75 NGFR-I) regions to same age untreated and preimmune sera-treated littermates. We confirm previously reported localizations of p75 NGFR-I in the rat brain. In addition, we demonstrate that p75 NGFR-I levels of ANTI-NGF-treated rats found in the molecular, the granular and the Purkinje cell layers of the cerebellum, the vestibular nuclei, the spinal tract of V and the cochlear nuclei remain at lower concentrations compared to same-age control animals. We also demonstrate that p75 NGFR-I levels in the basal nucleus approaches background levels after ANTI-NGF treatment. We hypothesize that ANTI-NGF biologically inactivates NGF, which over a period of 30 days results in decreased p75 NGFR-I. These results are consistent with neuronal loss in these regions following ANTI-NGF treatment. Furthermore, the immunological methods used to produce the specific deficits in the present study may have broader implications with respect to usefulness as a method for determining the dependency of CNS neuronal populations for a putative neurotrophic factor and as a method for the development of models of neurodegenerative diseases.


Effect of chronic treatment with recombinant interleukin-2 on the central nervous system of adult and old mice.

We have studied the effects of chronic treatment with recombinant interleukin-2 on the central nervous system of adult and old mice. Treatment with high doses of recombinant interleukin-2, on a schedule similar to that used in humans, was started at the age of 4 and 17 months, respectively, and ended 3 months later. At that time, all the mice were tested for acquisition of a passive-avoidance task and then sacrificed for histological examination. Three of the four groups (treated and control adults and control old mice) did not differ from one another in task performance or neuron density in frontal cortex, cerebellum, dentate gyrus or CA1-2, CA3, CA4 hippocampal areas. The old treated mice were unique in showing impairment of the mnesic functions and marked neuronal cell loss and degenerative changes limited to the hippocampal regions. Immunohistochemical studies did not show any significant amount of immunoglobulins in affected areas. Our results suggest that in old mice the impairment of the mnesic functions after recombinant interleukin-2 administration is due to hippocampal neuronal damage.


The physiological effects of serotonin are mediated by the 5HT1A receptor in the cat's cerebellar cortex.

Serotonin is present in a fine beaded plexus in the cerebellar cortex of several mammalian species. In the cat, serotoninergic afferents arise from neurons located within the lateral, paramedian and peri-olivary reticular nuclei (Kerr and Bishop, J. Comp. Neurol., 304 (1991) 502-515). In addition to serotoninergic afferents, these same nuclei also contain a separate population of neurons that give rise to mossy fibers to the cerebellar cortex. Physiological studies have shown that mossy fibers are excitatory to their target neurons. The intent of the present study was to determine the physiological effects of serotonin in the cat's cerebellum in an in vivo preparation and to identify the receptor(s) that mediate the observed responses. Iontophoretic application of serotonin (5HT) onto Purkinje cells reduces the spontaneous firing rate of all cells tested (n = 12). Serotonin also blocks the excitatory effects elicited by the application of aspartate in 17 of 19 units tested and of glutamate (n = 62) in all cases. In addition, 5HT potentiated the inhibitory action of GABA (n = 12). Iontophoretic application of the 5HT1A agonists, 8-OH-DPAT and ipsapirone, mimic the suppressive action of serotonin in a dose-dependent manner. This response, as well as the 5HT mediated suppression are blocked by the application of spiperone, a 5HT1A antagonist. Compounds selective for the 5HT1C,2 and 3 receptors are physiologically ineffective. The present data are in partial agreement with previous studies in the rat's cerebellar cortex.(ABSTRACT TRUNCATED AT 250 WORDS)


bFGF promotes the survival of entorhinal layer II neurons after perforant path axotomy.

Infusion of basic fibroblast growth factor (bFGF) prevents the loss of cholinergic neurons in the septum/diagonal band of broca following fimbria-fornix transection. However, an in vivo test of whether bFGF will also rescue injured non-cholinergic or cortical neurons has not been carried out. Previous studies have shown that the majority of layer II stellate neurons utilize an excitatory amino acid as their neurotransmitter. In order to determine if bFGF acts on non-cholinergic cortical neurons, a paradigm was developed to examine whether or not bFGF could spare layer II entorhinal stellate cells from axotomy induced death or atrophy. Axotomy of the medial entorhinal cortex fibers projecting to the dentate gyrus of the hippocampal formation via the perforant path lead to retrograde cell loss in entorhinal cortex. Fourteen or thirty days after a unilateral knife cut axotomy of the perforant path, layer II of medical entorhinal cortex showed a 28% decrease in large stellate neurons as well as many weakly stained, hollow cells compared to the non-lesioned side or naive controls. Layer IV neurons, however, which do not project via the perforant path, showed little detectable change in the number of cells ipsilateral to the knife-cut as compared to the contralateral side. Intraventricular infusion of bFGF over a period of 14 days reduced the 28% cell loss to less than 6%. Thus, bFGF is capable of preventing cortical neuronal loss and/or atrophy associated with retrograde degeneration of non-cholinergic neurons following axotomy.


Effect of starvation on glycogen and glucose metabolism in different areas of the rat brain.

We have studied the changes in concentration of glycogen, glucose and the bisphosphorylated sugars, glucose 1,6-P2 and fructose 2,6-P2, in several rat brain regions during 72 h of starvation. The animals were killed by focused microwave irradiation. The activities of glycogen metabolizing enzymes in the different areas were measured. A large decrease in glycogen and glucose concentration was observed in all areas. The concentrations of bisphosphorylated sugars changed, suggesting that an increase in glycolysis could take place at the beginning of starvation, with blood glucose as a major energy source. Differences in metabolite concentration before starvation disappeared after 72 h. The activities of glycogen synthase, glycogen phosphorylase and glycogen phosphorylase kinase were similar in all areas, and they did not change during starvation.


Identification of a sexually dimorphic neural population immunoreactive for calcitonin gene-related peptide (CGRP) in the rat medial preoptic area.

The medial preoptic area (MPOA) of the rat exhibits morphological sex differences and is implicated in sex-specific functioning and behaviour. Using immunocytochemistry, the distribution and numbers of cells containing calcitonin gene-related peptide (CGRP) were examined in the MPOA of adult male and female rats. In the intact female rat, CGRP-immunoreactive (-IR) cells were found in a continuum within the MPOA extending from the caudal aspects of the organum vasculosum of the lamina terminalis through the anteroventral periventricular nucleus (AVPv) to the region of the medial preoptic nucleus (MPN). An additional small group of CGRP-IR cells was noted at the level of the caudal MPNin the ventrolateral (VL) region. Compared with males, the AVPv and MPN regions of the female contained over 25-fold more CGRP-IR cells (P < 0.01). The VL region contained similar numbers of CGRP-IR cells in both sexes. Ovariectomy 1 month earlier, with or without subsequent 17-beta estradiol treatment, had no effect on the numbers or distribution of CGRP-IR cells in the MPOA. Gonadectomy of male rats resulted in a significant increase (P < 0.01) in the numbers of CGRP-IR cells in the AVPv and MPN regions. Subsequent administration of testosterone propionate for 1 week reduced (P < 0.05) numbers of CGRP-IR cells to levels observed in the intact male. Neurones containing CGRP in the VL group were not altered by gonadal steroid manipulation. This study shows that CGRP neurones in the AVPv/MPN region are sexually dimorphic.(ABSTRACT TRUNCATED AT 250 WORDS)


Lesions of the medial septum which produce deficits in working/spatial memory do not impair long-term potentiation in the CA3 region of the rat hippocampus in vivo.

The effects of removing the septohippocampal pathway on the ability to induce long-term potentiation (LTP) in the CA3 region of the hippocampus was examined in vivo in rats. The septal input to the hippocampus was destroyed by electrolytic lesioning of the medial septum (MS). Prior to electrophysiological investigation, working/spatial memory of lesioned and control rats was tested using an 8-arm radial maze task. Maze performance was significantly impaired in animals with MS lesions. LTP inducibility was examined in the commissural fimbrial fibre- and mossy fibre (mf)-CA3 pathways in MS-lesioned and control rats. The pre-tetanus values in MS-lesioned rats tended to be smaller than those in controls, in both pathways. High-frequency stimulation of the commissural fibres resulted in a sustained increase in the orthodromic population spike and EPSP amplitude in both control and MS-lesioned rats. The magnitude of potentiation was similar in both groups. In control rats, high-frequency stimulation of the mf potentiated the amplitude of both the population spike and EPSP; in MS-lesioned rats, the EPSP amplitude alone was significantly increased by mf high-frequency stimulation. Hippocampal acetylcholinesterase (AChE) content was severely reduced bilaterally in MS-lesioned rats with working/spatial memory impairments, indicating that the lesions were effective in destroying the cholinergic septohippocampal input. These findings suggest that, in contrast to working/spatial memory processes, LTP at CA3 synapses is not dependent upon the integrity of the septohippocampal pathway.


Plasticity of GABAergic terminals in Deiters' nucleus of weaver mutant and normal mice: a quantitative light microscopic study.

This study reports on the developmental changes in size and the average density of GABAergic axonal boutons bordering on the somata of large neurons in the dorsal part of the lateral vestibular nucleus (Deiters' nucleus) in normal and mutant mice. Weaver mutants, PCD mutants and the corresponding wild types were used to test for size alterations and differences in the number of GABA-immunopositive terminals. Hemicerebellectomized animals were examined in addition. Quantification of bouton profile size was performed from 30-microns-thick vibratome and 0.5-micron Araldite-embedded semi-thin sections immunoreacted for GABA from 7 days postnatally up to an age of 9 months. Terminal density was determined at the 5-6 month stage from semi-thin sections only. Morphometric analysis over the lifetime of normal animals (B6CBA) revealed a progressive increase in the size of bouton profiles, which peaked at 5-6 months and reached sizes of 2-3 microns2. In weaver mutants a parallel development in terminal size was found to be present, but the size of the largest terminals exceeded those of the controls by 75-100%, reaching 3-6 microns2 with the same time course. PCD mutants, with an almost total absence of Purkinje cells had, on the contrary, small bouton profiles that reached a maximum of only 2 microns2. The hemicerebellectomized animals responded with decreased bouton profile size ipsilaterally. The terminal numbers per unit membrane length were surprisingly similar in wild types and weaver mutants, despite a reduction in Purkinje cells of almost 50% in the weaver anterior lobe.(ABSTRACT TRUNCATED AT 250 WORDS)


Cortical neurons inhibit basal and interleukin-1-stimulated astroglial cell secretion of nerve growth factor.

Primary cultures of neonatal rat cortical neurons and astrocytes synthesize and secrete nerve growth factor (NGF). Co-culturing neurons with astrocytes decreased NGF secretion in the co-cultures. The inhibition of co-culture NGF secretion was partially reversible upon selectively decreasing the number of neurons by glutamate treatment. Interleukin-1 beta (IL-1) stimulated NGF secretion from astrocytes, and the magnitude of this secretion was decreased in the co-cultures. Thus, co-culture with neurons decreases astroglial cell secretion of NGF and down-regulates astroglial responsiveness to IL-1.


Octopamine selectively modifies the slow component of sensory adaptation in an insect mechanoreceptor.

The effects of octopamine were studied on the dynamic behavior of the sensory neuron in the cockroach femoral tactile spine. The neuron is a rapidly adapting mechanoreceptor in which adaptation occurs by elevation of the threshold for action potential encoding. The threshold follows increases or decreases of membrane potential, with a delay that involves two separate exponential components. Previous evidence has associated the slow component with sodium pumping and the fast component with sodium channel inactivation. Octopamine reversibly raised the resting threshold and increased but slowed the slow component. These data indicate that octopamine has specific effects on membrane-ionic processes in insect sensory neurons.


Dynamic analysis of drug action on in vitro reconstituted thyroid follicle by microinjection of tracer molecules and videomicroscopy.

Thyroid cells isolated from the gland by trypsinization are capable in culture of reconstituting histiotypic structures, the thyroid follicles. This morphological differentiation requires the presence of the main thyroid regulator; thyrotropin. We have analyzed some structural and functional aspects of in vitro reconstituted thyroid follicles (RTF) using microinjection of fluorescent probes and videomicroscopy. This experimental approach allowed to visualize biological processes and actions of drugs, signalling factors, etc. in living cells. We describe here some examples of what can be studied with this powerful still-undervalued method. Microinjection of a cell-impermeant fluorescent probe of either high or low molecular mass into the lumen of RTF allowed to check the tightness of this compartment and therefore to analyze the control of tight junctions assembly. A small cell-impermeant probe like Lucifer Yellow microinjected into a cell was used to demonstrate and then to study the regulation of cell to cell communication via gap junctions. The presence of calcium in the lumen of RTF was detected by microinjection of a properly designed probe: Calcium Green which becomes fluorescent in the presence of the ligand. The lumen to cell transport or endocytosis of thyroglobulin, the thyroid prohormone, which is stored into the lumen of the follicles, is currently studied by microinjection of TRITC-labeled thyroglobulin. Coupled to image processing and videorecorder systems, kinetic analysis and quantitative measurements can be performed.


Tracheal epithelium in culture: a model for toxicity testing of inhaled molecules.

Rabbit trachea primary cultures have been developed as a model to evaluate the toxicity of noxious airborne pollutants. A mucociliary epithelium has been restored in vitro on collagen gel. Several general cytotoxicity assays (viability and growth inhibition) permit a first assessment for the acute toxicity of the tested molecules. More specific criteria such as measurement of the integrity of the epithelial barrier and inhibition of ciliary beat frequency allow to determine a specific impact of xenobiotics on the mucociliary epithelium in culture.


Use of mesothelial cell cultures to assess the carcinogenic potency of mineral or man made fibers.

Natural mineral fibers may produce pulmonary cancers and mesothelioma. In contrast with lung cancer, the incidence of fiber-induced mesothelioma is not enhanced in smokers compared to non smokers. It is therefore of special interest to use mesothelial cells to study the toxicity of natural or man made mineral fibers. Several years ago, we have developed a method to culture rat pleural mesothelial cells (RPMC). We have first studied the effects of asbestos fibers by the application of in vitro tests formerly developed to determine the genotoxicity and transforming potency of soluble xenobiotics. Moreover, we have determined whether RPMC expressed cytochromes P450 known to metabolize polycyclic aromatic hydrocarbons. This paper reviews the results obtained so far. It has been found that asbestos fibers produced a cell transformation and a genotoxicity characterized by the formation of aneuploid cells, abnormal anaphases, chromosomal aberrations and DNA repair (UDS). In addition, RPMC expressed different forms of cytochromes P450. It is nowadays suggested that the tumorigenic potency of asbestos fibers may be related to the fiber dimensions, to their surface properties and in vivo biopersistence; this term involves the fiber solubility in biological medium and the fiber epuration from the lung by clearance mechanisms. Experiments are now in progress to determine whether the in vitro effects are dependent on the fiber parameters suggested as playing a role in the carcinogenic potency.


In vitro study of gas effects on alveolar macrophages.

To evaluate the biological effects of gas pollutants on alveolar macrophages several in vitro systems have been developed. We described here an original method of cell culture in aerobiosis, which permitted direct contact between the atmosphere and the target cells. We studied the long term (24 h) and short term (30 min) effects of NO2 on alveolar macrophages. Our results demonstrated that exposure of alveolar macrophages to gas pollutants may be responsible for either cell injury or cell activation associated with the release of various bioactive mediators (superoxide anion, neutrophil chemotactic activity). Cell culture in aerobiosis opens new ways for the research on the biological effects of gas pollutants.


Dual compartment (bicameral) culture: role of basement membrane in epithelial differentiation.

A number of years ago we reported that tight junctions between adjacent Sertoli cells subdivide the seminiferous epithelium into two compartments, basal and adluminal, thus forming the morphological basis of the blood-testis barrier. It is now generally believed that the special milieu created by the Sertoli cells in the adluminal compartment is essential for germ cell differentiation. In order to duplicate the compartmentalization that occurs in vivo, Sertoli cells were cultured in bicameral chambers on Millipore filters impregnated with a reconstituted basement membrane. Confluent monolayers of these cells were tall columnar (40-60 microns in height) and highly polarized. These Sertoli cell monolayers established electrical resistance that peaked when the Sertoli-Sertoli tight junctions developed in culture. In addition, the monolayers formed a permeability barrier to 3H-inulin and lanthanum nitrate. The bicameral chambers were utilized in a number of studies on protein secretion, and it was revealed that numerous proteins are secreted in a polarized manner. In another study, hormone-stimulated aromatase activity was measured in Sertoli cells grown on plastic culture dishes, plastic dishes coated with laminin or Matrigel, and in the bicameral chambers. Cell culture on basement membrane substrates decreased the FSH-dependent estrogen production. No estrogen production was observed when the Sertoli cells were cultured in the bicameral chambers. These results are in accord with the hypothesis that differentiated Sertoli cells lose their ability to metabolize androgen to estrogen in an hormone-dependent manner, whereas undifferentiated cells in culture, or in vivo, have a very active FSH-dependent aromatase activity.(ABSTRACT TRUNCATED AT 250 WORDS)


Genetic influences on reproductive performance.

The potential for genetic improvement of reproductive performance in pigs is great. Improvement can be achieved by increased commercial use of F1 hybrid females produced by crossing lines with excellent reproductive performance and by selection within nucleus populations of these lines for improved reproduction as well a for growth and carcass characteristics. The value of incorporating reproductive traits in selection programs is critically dependent on the relative economic values of fat (or lean) and of extra pigs produced. There is widespread agreement that litter size is the first choice as both the selection objective and the criterion to improve reproductive performance, on economic, genetic, and ease of measurement grounds. Although there are few examples of successful response to selection for increased litter size in pigs (exceptions being the University of Nebraska selection experiment and results achieved by hyperprolific selection schemes), overall results do not suggest that selection for litter size is impossible, only that it is difficult with limited resources. Artificial insemination is being used increasingly by pig breeding companies to accelerate rates of genetic improvement and to improve dissemination. Combining data from genetically connected herds and analyzing with BLUP computer programs to make efficient use of information on relatives makes selection for litter size feasible while maintaining selection pressure for growth and carcass traits. Although selection for lifetime productivity is impractical today, the knowledge and data accumulated from successful breeding programs for litter size may help make it the selection objective of the future. Meanwhile, the search continues for useful indirect criteria for selection, from testis size to molecular markers, and scientists are working with highly prolific breeds of Chinese pigs to better understand the physiologic and genetic basis of large litter size.


Permeable collagen-glycosaminoglycan cross-linked copolymers for the study of biological responses of cocultured Sertoli and spermatogenic cells.

A novel collagen-glycosaminoglycan (C-GAG) substrate was developed to overcome the optical opacity of a HATF nitrocellulose substrate and to provide a more physiological permeable substrate for cocultured Sertoli and spermatogenic cells. Cocultures were prepared on optically transparent C-GAG discs attached to a polyester mesh to facilitate handling. Sertoli cells displayed a cuboidal-to-columnar shape; a large number of spermatogonia and primary spermatocytes connected by intercellular bridges were associated with basolateral and apical surfaces of Sertoli cells up to 12 days after plating. Rat Sertoli-spermatogenic cell cocultures have been used for testing the effect of toxicants on rat spermatogenesis in vitro. In our initial studies, we tested the effects of the toxicant gossypol on spermatogenic cells cocultured with Sertoli cells on non-permeable (plastic) and permeable substrates (HATF nitrocellulose) under both standard culture conditions and during perifusion after achieving a continuous electrical-resistant cell monolayer. A selective mitochondrial structural damage was observed in spermatogenic cells (spermatogonia and spermatocytes) but not in the coexisting Sertoli cells. This damage was time- (15-60 min) and dose-dependent (0.1-10 microM) and developed more rapidly under perifusion conditions. Similar mitochondrial damage was reported in the intact animal but required higher concentrations (mg) and longer administration time (months) for detection. Studies are in progress to evaluate the effect of additional toxic chemical agents on functional properties of Sertoli and spermatogenic cells in cocultures prepared on various classes of C-GAG substrates.


The Sertoli cell in vivo and in vitro.

The Sertoli cell extends from the basement membrane of the seminiferous tubule towards its lumen; it sends cytoplasmic processes which envelop different generations of germ cells. The use of Sertoli cell culture began to develop in 1975. To reduce germ cell contamination immature animals are generally used as Sertoli cell donors. Sertoli cell mitosis essentially occurs in sexually immature testes in mammals; mitosis of these cells is observed in vitro during a limited period of time. Sertoli cells in vivo perform an impressive range of functions: structural support of the seminiferous epithelium, displacement of germ cells and release of sperm; formation of the Sertoli cell blood-testis barrier; secretion of factors and nutrition of germ cells; phagocytosis of degenerating germ cells and of germ cell materials. Some of the Sertoli cell functions can be studied in vitro. The recent development of Sertoli cell culture on permeable supports (with or without extracellular matrix) has resulted in progress in understanding the vectorial secretion of several Sertoli cell markers. In addition to FSH and testosterone, several other humoral factors are known to influence Sertoli cell function. Furthermore, myoid cells bordering the tubules as well as germ cells are capable of regulating Sertoli cell activity. Sertoli cells are the most widely used testicular cells for in vitro toxicology. The testis is highly vulnerable to xenobiotics and radiations, yet the number of studies undertaken in this field is insufficient and should be drastically increased.


Biosecurity and minimal disease herds.

The minimal disease concept is a way of raising pigs so that some specific diseases are absent. Many bacteria and viruses can be transferred by pigs, air, or mechanical contact. To avoid contamination, the herd location should take into consideration disease transmission possibilities. Herd health status and source herd health status should be continuously monitored. To maintain herd health status, specific rules need to be followed for herd construction and establishment, compound perimeter, people movement, down time, animal transportation, feed use and delivery, vehicle movement, material, dead animal disposition, and rodent control. All new incoming animals should go through quarantine, and in some herds, safer methods such as AI, embryo transfer, MEW, or hysterectomy and fostering need to be used.


Breeding and gestation facilities for swine. Matching biology to facility design.

The rate-limiting phase of commercial swine production is breeding. The design of the breeding/gestation facilities influences the capacity of the producer to efficiently manage the breeding stock around the time of service and during the postservice period. Thus, breeding design potentially affects total born litter size and farrowing rates. In general, with original construction and facility renovation, the three principal activities of the breeding/gestation phase of production, (1) boar exposure/estrus detection, (2) breeding, and (3) gestation and pregnancy detection, should be independently considered.


Hormonogenesis in thyroid cells cultured on porous bottom chambers.

Different processes implied in thyroid hormonogenesis (thyroglobulin, thyroperoxidase and hydrogen peroxide generating system expressions) and their regulation by TSH and iodide have been studied using porcine thyroid cells cultured in porous bottomed chambers. This system allowed to reproduce the functional bipolarity. Cells form a tight and polarized monolayer. Both apical and basolateral poles of epithelial cells were independently accessible and the cell layer separated two compartments which can contain different media. A major polarized secretion of thyroglobulin into the apical compartment was observed; it was increased in the presence of TSH as well as the thyroglobulin synthesis and mRNA level. These TSH effects were the consequence of adenylcyclase stimulation. Active transport of iodide, iodination of thyroglobulin and hormonosynthesis took place only in the presence of TSH. These steps occurred at the apical pole of cells. In the culture chamber system, thyroglobulin was weakly iodinated (6 atoms of iodide per mole of thyroglobulin; in vivo up to 40 atoms per mole) but hormonogenesis efficiency was close to this one observed in vivo (40%). Iodide concentrations higher than 0.5 microM daily added to the basal medium inhibited iodination of thyroglobulin and hormonosynthesis. Some components contained in culture media were inhibitors for iodination when they were present in the apical medium such as vitamins, amino acids and phenol red. The culture system appears to be interesting for pharmacological and toxicological studies.


Factors affecting the reproductive performance of the weaned sow.

The majority of sows return to estrus within 2 weeks after weaning. Swine practitioners attempt to optimize production by reducing the WEI. Some sows fail to resume estrous cyclicity after weaning; however, the endocrinologic pathogenesis of these anestrous sows is speculative. The average WEI is influenced by numerous factors, including season, environmental temperature, photoperiod, nutrition, stress, facility design, lactation length, and management practices. It is evident that the majority of these factors have a more profound influence on primiparous sows than on multiparous sows. Optimum protein and energy consumption by sows during lactation and after weaning and effective utilization of breeding facilities reduce the WEI. The precise roles of photoperiodic changes, elevated environmental temperatures, and stress in seasonal infertility remain poorly understood. Fortunately, current management techniques have reduced the WEI on most farms without instituting therapeutic measures.


Mating management.

The mating management of pigs is conducted under a wide spectrum of management techniques because females are variously hand-mated, pen-mated, or artificially inseminated. Factors that may affect the onset of estrus when hand-mating are boar exposure, method of housing females after weaning, season, type of facilities, and parity. The expression of sexual behavior in both male and female pigs can be influenced by a variety of physiologic, psychological, and environmental factors. To optimize the efficiency of estrous detection, females should not receive boar stimuli just prior to estrous detection when either hand-mated or artificially inseminated. When females are pen-mated, management procedures need to be employed to prevent an excessive number of estrous females from accumulating, which leads to a decrease in boar fertility.


Artificial insemination in swine.

This article analyses the advantages and disadvantages of artificial insemination with semen purchased from a center as well as from the herd boars on the farm. Intensive swine production could benefit greatly by adapting artificial insemination with herd boars, particularly from savings in labor and boar numbers. The techniques for semen collection, extension, and insemination are described, and sources for equipment given. Expected results of artificial insemination are quoted from experiments and international field experience.


Optimizing longevity in sows and boars.

Longevity of sows and boars affects the productivity and economics of the herd. Implement a culling program that is tailored to the specific needs of the producer and the characteristics of the farm. Evaluate the removal policies regularly, estimate the longevity of breeding animals, calculate the nonproductive sow days associated with culling, and determine the major causes of culling and death. Once a longevity problem and its extent have been identified, the underlying problems and predisposing factors can be more easily corrected.


Managing the lactating sow for optimal weaning and rebreeding performance.

Management of the lactating sow influences milk production and subsequent reproduction through changes in nutrient intake. The management goal during lactation is to maximize feed intake. Decreasing the effective environmental temperature, increasing the nutrient density of the lactation diet, maintaining fresh adequate supplies of feed and water, and preventing excess weight gain during the prior gestation period will increase nutrient intake during lactation. Effective environmental temperature of the lactating sow can be maintained in the thermoneutral zone by using drip cooling, increased ventilation rates and flooring materials with superior conductive properties. Sow parity, genetics, litter size, and disease level will also influence feed intake. Management practices must account for these factors and, thus, should be tailored to individual farm situations to ensure adequate nutrient intake and prevent aberrations in subsequent reproductive performance.


Management of replacement breeding animals.

In conclusion, replacement gilts should be finally selected before 110 kg body weight and should have been stimulated to reach puberty by this time. Currently, the best method for achieving this is ad libitum feeding and the correct use of boar exposure. However, whether ad libitum feeding will always be the best feeding strategy remains to be seen. The impressive success of selection programs and the potential for the use of growth promoters and repartitioning agents in the feeder barn may require that future replacements be identified at an early stage and raised under a separate feeding regimen. To produce a large first litter, gilts should be bred at their second estrus. There is unlikely to be any further increase in litter size by delaying mating to third estrus. A major consideration when choosing when to breed replacements is the length of their productive life. For many farms, this may be maximized by delaying breeding until the sow is 120 to 130 kg body weight with a minimum backfat depth of 18 mm. For the producer who is able to exert control over lactation condition loss, however, breeding replacements at about 110 kg body weight with backfat depths of 14 to 16 mm should not adversely affect long-term performance. Although it is true that, under some conditions, the lean gilt may become a problem, it is equally true that her potential is as great as her fatter contemporaries. Whether this potential is realized is largely under the producer's control. Regarding the boar, nutritional management need not vary from that provided for gilts. However, if replacement boars are raised on-farm, it is of great importance that they be allowed to socially interact with other pigs. Failure to provide this opportunity will result in a boar with reduced libido.


Nutrition for optimizing breeding herd performance.

Modern sows are younger and leaner at time of mating and probably have poorer appetites than sows of 10 to 15 years ago. Therefore, feeding strategies should aim to minimize weight loss and maintain a sow's body condition throughout her reproductive life. The efficiency with which gilts are introduced into the breeding herd is as important in economic terms as is the efficiency with which the sow returns to estrus after weaning. Gilts should be selected at 50 to 60 kg, and fed a 16% protein diet ad libitum until mated at their second estrus, when they weigh 115 to 120 kg and have 17 to 20 mm backfat. Flushing gilts before the onset of second or third estrus increases ovulation rate of restricted gilts to the levels achieved by gilts fed ad libitum. During gestation, maintenance represents 75 to 85% of total energy requirements. The aim should be to achieve 20 to 25 mm backfat at farrowing. Increased feed intake from day 2 to 3 after mating will not increase embryo mortality. Feeding an extra 1 kg feed/sow/day for the last 10 days of gestation increases piglet birth weight slightly and prevents a loss of 1.5 to 2.0 mm of sow backfat. Wherever possible, sows should be fed ad libitum from the day after farrowing until weaning. Reduced feed intake by lactating sows, for whatever reason, results in excessive weight and condition loss. Excessive weight loss in lactation causes extended remating intervals, a lower percentage of sows returning to estrus within 10 days of weaning, reduced pregnancy rate, and reduced embryo survival. Ovulation rate is not affected by level of feed intake in lactation. It has been suggested that sows will have minimum weaning-to-service intervals when they weigh 150 kg or more at weaning. It is likely that the sow must be anabolic for about 10 days before she will exhibit postweaning estrus. The decision when to rebreed is made some time prior to weaning and is mediated by a host of substrates, hormones, and neurotransmitters. Sows with a delayed return to estrus also have a lower pregnancy rate and smaller subsequent litters. If sows lose considerable weight or condition during lactation, a high level of feeding in the postweaning period will improve embryo survival.


Optimizing farrowing rate and litter size and minimizing nonproductive sow days.

The most universally accepted measure of reproductive performance is PSY. Excellence is achieved by reducing NPD, increasing liveborn litter size, and reducing preweaning mortality. To reduce NPD, farm management should be directed to improve farrowing rate. Matings that occur in late summer usually are less fertile as the result of the pig's normal seasonal variation. Confinement gestation stalls are the best prevention for the reduced farrowing rate caused by season. Liveborn litter size is the result of a mixture of genetic and management variables. F1 females have the highest heterosis. The management variables of age at first mating, wean-to-service interval, skip mating, lactation length, and parity all influence litter size.


Stillbirths, mummies, abortions, and early embryonic death.

Stillbirths, mummies, abortions, and early embryonic death have a substantial impact on the profitability of a farm in both endemic and epidemic conditions. Fetal death is highly dependent on stage of gestation. Implantation occurs around day 14 postmating in sows, and fetal death of an entire litter at this time usually results in a regular return to service. If more than four embryos remain alive, the sow may go on to farrow normally. If fetal death occurs after implantation but before calcification (around 35 days gestation), the sow will either return to estrus at an irregular interval or will farrow a normal litter of reduced size. Although fetuses are normally resorbed prior to calcification, fetal death after that stage of development leads to mummification. Abortions are more directly related to maternal control of pregnancy than fetal failure. Stillbirths are those pigs that appear normal at birth but have lungs that do not float in water. Causes of fetal death can be divided into infectious and noninfectious categories. Infectious causes perhaps are overemphasized but are certainly important in epidemic situations. Some infectious causes of fetal death are primarily systemic maternal pathogens, whereas others may attack the fetus and/or placenta, directly such as PPV, PEV, PRV, SIRS virus, and Leptospira sp. Several other infectious agents have been associated with fetal death. Noninfectious causes of stillborns, mummies, abortions, and early embryonic death are most common in endemic situations. Most stillbirths are due to difficulty at or around parturition, primarily extended duration causing fetal anoxia. Environmental factors such as increased ambient temperature and seasonal infertility affect death rates, as do specific individual sow characteristics, nutritional factors, and toxicities. The causes of stillborns, mummies, abortions, and early embryonic death are often difficult to ascertain, but the potential rewards make investigation efforts worthwhile.


Porcine urogenital disease.

Porcine urogenital disease is the result of an imbalance of the normal microflora of the urinary and reproductive tracts brought about by hormonal, environmental, and management-related stress factors. Production and economic losses can be substantial, and diagnosing and treating the problem can be frustrating. Through proper hygiene, facility design, and culling procedures, the severity of the problem can be minimized.


Pathogenesis, prevention, and treatment of lactational insufficiency in sows.

Lactational insufficiency is one of the major problems in swine production, and the consequences on the growth of the litter must be emphasized. Pathophysiology involves neuroendocrinologic interactions and the role of target organs such as the mammary glands, uterus, bladder, and gut. The complexity of these interactions and the fact that all interactions are not well understood leads to a broad approach. Some risk factors and their associations with lactational insufficiency are described to help the practitioner in the clinical approach to such a problem.


Preweaning mortality.

A number of factors, both infectious and noninfectious, contribute to preweaning mortality in pigs. It is important to establish the causes of preweaning mortality and their distribution in investigating farm problems. The various causes, risk factors, and a step-by-step approach to solving the problem are discussed.


Pharmacologic control of swine reproduction.

Current approaches to control fertile estrus and parturition in the pig are discussed. Techniques to induce estrus in acylic pigs (pregnant mare serum gonadotropin and human chorionic gonadotropin) to synchronize estrus in cyclic females (progestins), and to control onset of ovulation (human chorionic gonadotropin) are described. Regimens that induce parturition (prostaglandins) and precipitate farrowings at more predictable times (oxytocin) are addressed.


Financial evaluation and decision making in the swine breeding herd.

As the swine industry continues to evolve and develop a greater business orientation--one that demands justification of costs, benefits, and risk-exposure--the veterinarian will be confronted with the need to develop an economic basis to supplement the biologic. The profession should view such a challenge as an opportunity to broaden its role in serving the businesses that make up the industry. However, to be effective in such an expanded role requires that the veterinarian develop a good working knowledge of risk and risk management as well as the role played by economic evaluation techniques used in financial evaluation in the risk management process. Several potential advantages exist for the veterinarian who determines to expand his or her risk management role by using financial evaluation in the course of day-to-day practice routine: Additional service or consulting income may be generated through evaluating alternatives through decision analysis. Client confidence may be strengthened in the veterinarian and in the recommendations made. The costs of consulting services and health products for the client may be viewed as income-generators instead of being viewed solely as expenses. Referrals of additional progressive clients may be generated by satisfied current clients. The veterinarian may achieve personal satisfaction through substantiating his or her recommendation beyond clinical or experimental judgment. With the capability of using financial evaluation techniques in the evaluation and justification of recommendations comes significant responsibility. Reliance upon financial evaluation techniques as a more objective approach in the management of risk for clients carries with it an inherently greater degree of trust. Such trust must be deserved. The universal adage "garbage in, garbage out" applies directly to financial evaluation. Financial evaluation techniques have the potential to provide the decision-maker with objective and accurate analysis, improving the probability of choosing an optimal course of action for the business. However, the objectivity and accuracy achievable is highly dependent upon the quality and quantity of information used in the analysis. Veterinarians must be cognizant of the increased responsibility demand placed on those using financial evaluation techniques to direct the course of action for their clients. Financial evaluation tools are available to decision-makers that may improve the objectivity of the decision-making process, providing for a greater degree of confidence in the expected outcome. Stated another way, these tools can reduce the number of surprises a decision-maker encounters as a result of the decisions made.(ABSTRACT TRUNCATED AT 400 WORDS)


[Experiences of mammaplasty in 32 patients of macromastia and mammary descensus].

32 patients with macromastia and mammary descensus have been treated since 1982 by means of mammoplasty. The Mckissock's operation was modified to make the surgical procedure simple and rational, easy to grasp, safe and reliable. According to our experiences, the advantages of the operation, the complications and their prevention and management have been discussed in the present communication.


[Clinical analysis of fluid resuscitation in severe burned patients with or without inhalation injury].

The amount of fluid resuscitation, estimated according to crystal-colloid for formula, were retrospectively studied in 55 cases of severe burned patients with or without inhalation injury. These cases were divided into two groups. One group consisted of patients with inhalation injury, while the another there was no inhalation injury. The result showed that patients in the former required 13.5% additional amount of fluids over the latter in the first 24 hours postburn, and this amount was mainly due to the additional requirement in the first 8 hour period. We propose that the amount required in severe burned patients with inhalation injury in the first 24 hours postburn should be appropriately increased, and special attention should be paid to the harmful effect of inadequate fluid resuscitation in the first 8 hour period. The amount of fluids required for resuscitation in severe burned patients with inhalation injury, as estimated according to crystal-colloid formula, should be increased by 13.5% in the first 24 hours, and 2/3 of the addition amount should be given in the first 8 hours, with 1/3 of it in the second and third 8 hours postburn.


[Transplantation of thin pedicle skin flap: a new method in the treatment of skin defect of hand (a report of 54 cases)].

We used the transplantation of the thin pedicle skin flap to repair the skin defect on the hand and claw hand due to scar contracture in 54 patients. The results were improved. The size of the skin flaps was 16-8 cm x 6-4 cm. The average time of division of the pedicle was 10.5 days. In comparison with the free skin graft with a preserved subcutaneous vascular network, the whole thickness free skin graft, the random pattern skin flap (local flap, fascio cutaneous flap and traditional skin flap), the axial pattern skin flap (including island skin flap and myocutaneous flap), showed that the thin pedicle skin flap was superior over them. The indications, blood circulation and the points for attention in the operation were discussed.


[An experimental study of mitogen-stimulated blastogenic transformation and interleukin-2 in mice after burn injury].

In this study, mitogen-stimulated blastogenic transformation (MSBT) and interleukin 2 (IL-2) production by splenic lymphocytes in mice were measured at various time intervals after unanesthetized burn injury. The results showed that both MSBT and IL-2 production were suppressed after burn injury. There was a significant positive correlation between these two parameters. The postburn serum showed in vitro suppressive activity upon MSBT, IL-2 production and IL-2-IL-2R interaction of normal control. The results indicated that burn injury had a significant effect on lymphocytes.


[Changes in malondialdehyde contents in serum and tissues after ischemia and reperfusion of the bowel in dogs].

The portal circulation was reduced to 50% for one hour by partially occluding the superior mesenteric artery, and the relationship between the changes in malondialdehyde (MDA) and multiple organ damage was studied. The results showed that there was a marked change in hepatic enzymology, with MDA values significantly elevated in serum and lung and liver tissues after the reperfusion when compared with controls. These results suggested that lipid peroxidation initiated by free oxygen radicals played an important role in the multiple organ damages, and the extent of organ damage was closely correlated with the contents of MDA in tissues.


[Uses in fibronectin on eschar-excised wound to promote expansion of skin grafts].

Fibronectin was applied to wounds resulted from excision of burn eschars at different time intervals after burn in piglets, for the purpose of studying the effect of Fn in promoting the expansion of skin grafts which were laid on the wounds. Fn content in wound tissue was measured by means of ELISA, while tissue Fn and Type-III collagen localization was identified by immunohistochemical method ABC. The rate of the graft expansion was then measured by Automated Scanning Image Analyzer. The results showed that topical application of exogenous Fn on the wound could rapidly increase the density gradient of tissue Fn on the wound and promote expansion of skin grafts.


[Effect of crystal No 4 of Polygonum cuspidatum on microcirculatory disturbances during burn shock].

A model of burn shock was reproduced in rat. The effects of crystal No 4 of Polygonum cuspidatum (p. c.) on microcirculation of spinotrapezius muscle in rat with burn shock were observed with a special Hitachi TV set with magnification of 4000x. The changes in water content in tissues of lung and burned skin were measured. The survival time of burned animals was recorded. The aggregation of WBC and the degree of tissue damage in lung were examined in pathologic slides. The results showed that during burn shock the number of adhesive WBC in venules was about 8 times more than normal, the amount of open capillaries was reduced by 3/5 of normal. There were apparent aggregation of WBC and tissue damage in the lung. The number of adhesive WBC was decreased by 80%, the amount of open capillary returned to near normal, and the aggregation of WBC and the degree of tissue damage in the lung were alleviated by administration of crystal No 4 of Polygonum cuspidatum. The survival time in p. c.-treated group was prolonged 1.9-fold of that in n.s.-treated group. The results indicated that the decrease of WBC adherence was caused by, on the one hand, the increase of driving force and wall stress after treatment of p.c., and on the other hand, p. c. might affect some humoral factors which promoted WBC adherence.


[Acceleration of epidermis proliferation by direct current stimulation (an experimental study)].

An experimental study was designed to evaluate the effect of direct current stimulation on the rate of epidermis proliferation using guinea pigs. In the study, two full thickness skin defects were made over the paravertebral region in each animal. The edges of wounds were transfixed. One of the wounds in each animal received the DC stimulation, and the other did not. The 44 animals were divided into four groups of eleven animals each, the stimulation strengths were 10 muADC, 30 muADC, 50 muADC, 0 muADC respectively. The results showed that the rate of epidermal cell proliferation and wound healing with the DC stimulation were faster, especially on the side of the negative electrode, and the rate of healing of the unstimulated wound in animals which received stimulation in the contralateral wound was faster than that of wounds of control animals.


[Plasmid fingerprinting technique analysis of Pseudomonas aeruginosa isolated from a burn unit].

We used plasmid fingerprinting technique to analyse 50 strains of Pseudomonas aeruginosa isolated from our burn unit and 15 strains of Pseudomonas aeruginosa from other wards. The results revealed that the carrier rate of R-plasmid was 100%. All 50 strains of Pseudomonas aeruginosa from burn units contained one plasmid with approximate molecular weight of 55 kb. Among 15 strains of Pseudomonas aeruginosa from other wards, 10 strains contained one plasmid with the same molecular weight. However, 3 strains were found to possess an additional 12.5-kd plasmid and 2 strains contained 77-kb plasmid. This proves that R-plasmid carrying Pseudomonas aeruginosa is prevalent in the burn unit, and R-plasmid is disseminated in the hospital.


[Study of the abnormal plasma proteins after burn injury].

In our previous study at least 4 abnormal bands were found in burned sera when analyzed by SDS-PAGE. In this paper further exploration of the characteristics of these abnormalities of burned sera were presented. By way of western blotting, 12 kinds of antisera against different human proteins as detecting antibodies were used. Results showed that bands developed by anti-haptoglobin antiserum corresponded well with the abnormal bands in SDS-PAGE. If the burned serum was separated by ion-exchange chromatography and its collected eluates containing protein peaks were tested for the immunosuppressive activities on normal murine spleen lymphocytes, the immunosuppressive eluates were usually found in the 6th protein peak. When these immunosuppressive eluates were further analyzed by SDS-PAGE and Western blotting, bands stained with anti-haptoglobin antiserum still coincided with the abnormal bands found in SDS-PAGE of burned serum. Result of this study suggests that one of the immunosuppressive factors found in burned serum may be a haptoglobin-like substance.


[Lengthening of residual digit by gradual traction and bone grafting (a report of 143 cases)].

Since 1985, a device for gradual traction was used for lengthening of 143 residual digits in 75 patients. The average lengthening was 2.2 cm. This method is simple, safe, and effective. The advantages of this method over the previously exercised tubed bone graft and other digital lengthening methods are that the digit reconstructed by the gradual traction-lengthening method has the normal cutaneous structure, good sensation function, and better contour. Multiple digits can be reconstructed simultaneously. Because the periosteum at the osteotomy site is kept intact, the long term results are consistent.


[Quantitative bacterial culture of subeschar tissue. A clinical study].

The relationship between the number of bacteria in the subeschar tissue and time, age, burn extent in 47 patients within a week after burn injury are presented. The results showed 1. There is a significant positive correlation between the number of bacteria in the subeschar tissue and time intervals following burn within the first week (r = 0.9839, P < 0.01); 2. In children (below 4 years) and patients over 45 years, there is a higher incidence of having bacterial counts of over 10(5)/gm of tissue than that of young and adolescent patients (P < 0.01); 3. There is a significant difference in subeschar bacterial count between major burns (> 30% TBSA) and smaller burns (< 30% TBSA P < 0.05); 4. Pseudomonas aeruginosa is one of the most frequent bacteria isolated, especially in major burns.


13C-CP/MAS NMR studies of the cyclomalto-oligosaccharide (cyclodextrin) hydrates.

The 13C-CP/MAS NMR spectra of cyclomaltohexaose (alpha-cyclodextrin) hexahydrate, cyclomaltoheptaose (beta-cyclodextrin) "undecahydrate", cyclomalto-octaose (gamma-cyclodextrin) "octadecahydrate", and of the same materials at lower levels of hydration are compared with solution NMR data, structures obtained from single crystal diffraction studies, and with previous reports of the 13C-CP/MAS NMR spectra. The chemical shifts of the C-1 and C-4 resonances can be correlated with the conformation about the (1----4) linkage. The chemical shifts of the C-6 resonances are also sensitive to hydrogen-bonding interactions, as shown by the spectral changes on loss of water from the structures. The results suggest that, for resonances of carbon atoms close to a centre of significant conformational change, chemical shifts may be predicted on the basis of conformation alone, but for the resonances of more distant atoms, changes in chemical shift due to conformational change may be masked by the effects of alterations in the local environment.


A spacer-modified disaccharide as a photoaffinity reagent for the acceptor-binding area of bovine (1----4)-beta-D-galactosyltransferase: comparison of its acceptor properties with those of other 2-acetamido-2-deoxy-beta-D-glucopyranosides.

The spacer-modified disaccharide 1,10-di-O-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)-2-azi-1,10- decanediol (10) that mimics the biantennary core heptasaccharide of N-glycoproteins has been synthesised. Compound 10 is an excellent acceptor in galactosyltransferase-catalysed galactosylation by UDP-galactose, is superior (7-8-fold) to analogues that have only one GlcNAc unit, and is an efficient photoaffinity reagent for galactosyltransferase. In the presence of UDP-Gal, no photoaffinity labelling by 10 takes place, which agrees with the mechanism of galactosyltransferase action.


Spacer-modified trisaccharide glycosides that mimic the biantennary Asn-linked oligosaccharide acceptor of (1----4)-beta-D-galactosyltransferase and can be used as competitive inhibitors and for irreversible deactivation.

The biantennary spacer-modified trisaccharide glycoside methyl 3,6-di-O-(2-acetamido-2-deoxy-beta-D-glucopyranosyloxyethyl)-alpha -D- mannopyranoside (5) was synthesised and used together with several 2-acylamino-2-deoxy-D-glucose derivatives in competition experiments with beta-D-galactosyltransferase. Compound 5 was an acceptor substrate (KM 0.18 mM) comparable to the biantennary core heptasaccharide of glycoproteins (KM 0.13 mM). Replacing the N-acetyl group by other N-acyl groups did not alter the kinetic parameters significantly. When the N-acyl group was iodoacetyl, the compound was an irreversible inhibitor.


Conformational analysis of the anomeric forms of sophorose, laminarabiose, and cellobiose using MM3.

Relaxed-residue energy maps based on the MM3 force-field were computed for relative orientations of the pyranosyl rings of sophorose, laminarabiose, and cellobiose, respectively the (1----2)-beta-; (1----3)-beta-; and (1----4)-beta-linked D-glucosyl disaccharides. Sixteen starting conformations of the rotatable exocyclic side-groups were considered for each molecule. All of the energy surfaces have two intersecting low-energy troughs and illustrate the importance of exo-anomeric effects in determining disaccharide conformation. Local minima were found by relaxed minimization without restriction. The energy surfaces of these disaccharides are very similar to the energy surfaces of their corresponding 6-methyltetrahydropyran analogues. There is good agreement between disaccharide structures having minimal MM3 energy and those found by crystallography.


Structural characterization of a novel diglycosyl diacylglyceride glycolipid from Rhizobium trifolii ANU843.

A novel glycolipid was isolated by chloroform-methanol extraction of Rhizobium trifolii ANU843 cells. Compositional analysis, methylation studies, 1H NMR and spectroscopies led to the identification of a diglycosyl diacylglyceride: 1,2-di-O-acyl-3-O-[alpha-D-glucopyranosyl-(1----3)-O-alpha-D- mannopyranosyl]glycerol. Iso-hexadecanoic and anteiso-heptadecanoic acids were the predominant fatty acids esterifying the glyceryl moiety, but a microheterogeneity in fatty acid composition was found, resulting in at least five distinct molecular species of the glycolipid. Although widespread in plants, animals and Gram-positive bacteria, glycosyl glycerides have been seldom reported in Gram-negative bacteria and this work is the first evidence of their occurrence in the bacterial family Rhizobiaceae.


Artificial antigens. Synthetic carbohydrate haptens immobilized on crystalline bacterial surface layer glycoproteins.

The crystalline surface-layer glycoproteins of Clostridium thermohydrosulfuricum L111-69, Bacillus stearothermophilus NRS 2004/3a and Bacillus alvei CCM 2051 were used for immobilization of spacer-linked blood group A-trisaccharide (alpha GalNAc(1----3)[alpha Fuc(1----2)]beta Gal) and of the spacer-linked, tumor-associated T-disaccharide [beta Gal(1----3)alpha GalNAc]. The immobilization involved the glycan portions of surface-layer glycoproteins. Different activation methods were used, namely, periodate oxidation, or treatment with epichlorohydrin or divinyl sulfone, followed by coupling of the hapten under appropriate conditions. The resulting conjugates are useful for assessing the application potential of haptenated surface layer preparations as carrier/adjuvants for the induction of immunity to poorly immunogenic molecules.


Structural elucidation of the capsular polysaccharide of E. coli serotype K47.

The capsular polysaccharide from Escherichia coli K47 was investigated using mainly methylation analysis and 1H and 13C NMR spectroscopy and shown to have the following repeating unit: [Formula: see text]


Structural variability in the glucuronoxylomannan of Cryptococcus neoformans serotype A isolates determined by 13C NMR spectroscopy.

Cryptococcus neoformans, the etiologic agent of cryptococcal meningoencephalitis, produces glucuronoxylomannan (GXM) as the major capsule component. Purified GXMs obtained from eight serotype A isolates of C. neoformans were treated by ultrasonic irradiation and then O-deacetylated prior to their comprehensive chemical analysis by GLC, GLC-MS, and 13C NMR spectroscopy. The average xylose: mannose: glucuronic acid molar ratio of the eight isolates is 1.96 +/- 0.25: 3.00: 0.58 +/- 0.10. Methylation analyses and 13C NMR spectroscopy show a general structure for GXM that is comprised of a linear (1----3)-alpha-D-mannopyranan substituted with beta-D-GlcpA and with beta-D-Xylp at O-2. Variable quantities of unsubstituted (1----3)-alpha-D-Manp were observed between the eight isolates studied. In several isolates some of the (1----3)-alpha-D-Manp residues are disubstituted with beta-D-GlcpA at O-2 and with beta-D-Xylp at O-4; this type of substitution was not previously thought to occur in serotype A isolates. Heterogeneity, between isolates, in the disposition of the substituents along the mannopyranan backbone was revealed by 13C NMR spectroscopy. The eight isolates, and three isolates previously studied, were each assigned to one of four distinct groups based on the 13C NMR chemical shifts of the anomeric carbons. Six of the eleven isolates gave identical spectra (Group I). The six major anomeric resonances from Group I were assigned to specific glycosidic linkages present in GXM. The remaining five isolates gave more complex spectra that are indicative of additional linkages and comprise the remaining three groups. Three of these five isolates contain substantial amounts of linkages previously thought to be distinctive of serotypes B and C, i.e., Manp residues that are 4-O-glycosylated with beta-D-Xylp. Methylation analyses only predicted an average repeating unit, whereas 13C NMR spectroscopy demonstrated that GXM from each isolate may be categorized into four groups by the occurrence of distinct sequences of carbohydrate residues.