EEG-dependent modulation of response dynamics of cat dLGN relay cells and the contribution of corticogeniculate feedback.

Single unit recordings were made extracellularly from the dorsal lateral geniculate nucleus (dLGN) in the anaesthetized and paralysed cat. The impulse rates of phasic (peak) and tonic components of visual responses to stimulation of the receptive field center by a flashing spot were determined during different states of the EEG, during local cortical cooling and during micro-iontophoretic application of the excitatory amino acid receptor agonists, quisqualate (QUIS) and N-methyl-D-aspartate (NMDA). Typically, visual responses were phasic during low frequency/high amplitude EEG patterns, resembling slow wave sleep (SWS). During high frequency EEG patterns (non-SWS) visual responses of X- and Y-cells exhibited a prominent tonic response component. This tonic component could be clearly reduced during ipsilateral cortical cooling in the non-SWS state. QUIS or NMDA, applied in order to mimic corticofugal activity, augmented the tonic response component, most efficiently during SWS EEG. The effects did not differ significantly for X- and Y-cells. During non-SWS EEG Y-cells exhibited a tonic response component similar to X-cells, but because of their higher peak rates the responses of Y-cells were on average more phasic than those of X-cells. Our results indicate that state-dependent changes in CNS activity modulate dL'GN responsiveness in part via the corticogeniculate feedback, and that predominantly the tonic response component is modulated.


A selective sexually dimorphic response in the median eminence neuropeptide Y.

Recent studies show that neuropeptide Y (NPY) is widely distributed in the hypothalamus and that it stimulates luteinizing hormone-releasing hormone (LHRH) release from the medial basal hypothalamus (MBH) of male and female rats. The neuroendocrine factors that regulate NPY neurosecretion in two sexes are not well understood. We have previously observed that orchidectomy (orch) in male rats decreased and testosterone (T) replacement increased NPY levels selectively in the median eminence (ME), arcuate nucleus (ARC) and ventromedial nucleus (VMN) and the KCl-evoked in vitro release of NPY from the MBH was likewise decreased after orch and restored by testosterone replacement. Now we report that these NPY responses are different in female rats following removal of the ovaries. Ovariectomy (ovx) decreased NPY concentrations in the ARC and VMN and not in the ME. Estrogen replacement restored NPY concentrations in the ARC and VMN, however, in the ME NPY concentration decreased significantly. Further, the KCl-evoked NPY release in vitro from either the MBH or ME of intact and ovx rats was similar. In an additional concurrent comparative study in male and female rats, we observed that of the 6 microdissected hypothalamic sites, NPY levels were approximately 50% less in the ME of intact females than in those of intact male rats. Again, in male rats there was a significant decrease in NPY levels in the ME, ARC and VMN after orch, whereas in female rats, NPY levels in the ARC and VMN, but not in the ME decreased after ovx.(ABSTRACT TRUNCATED AT 250 WORDS)


Neural correlates of genetically determined and acquired color preferences in quail chicks.

The uptake of tritiated 2-deoxyglucose (2-DG) was examined in brains of artificially selected and unselected control quail, in relation to genetically and environmentally variable early approach preferences of a colored stimulus. The data indicated high labeling in the tectofugal and low labeling in the thalamofugal projection systems in birds exhibiting preference for the light wavelength characteristics of the stimulus in independently tested choice situations. The differential labeling was observed regardless whether the stimulus preference was genetic or environmental in origin (i.e. whether it was exhibited by artificially selected and experientially naive or by unselected and imprinted control chicks). Chicks responding to the stimulus after prior imprinting experiences also exhibited high labeling in the intermediate medial portions of the neostriatum and hyperstriatum ventrale.


C-fos immunoreactivity in the brain following unilateral electrical stimulation of the dorsal periaqueductal gray in freely moving rats.

C-fos immunoreactivity was used to reveal brain areas in which neurons were influenced by electrical stimulations applied to the dorsal periaqueductal gray. These stimulations were applied in freely moving rats so that the resulting behaviors could be observed. Shortly afterwards, the brains of the rats were processed for C-fos immunoreactivity. In order to determine the specificity of the brain areas thus labeled, control stimulations were applied to the ventral tegmental area of other rats. Immunoreactive cells were found surrounding the tip of the stimulation electrode within a radius of 0.5 mm. This labeled area extended further along the rostro-caudal axis than along the medio-lateral or dorso-ventral axis in the periaqueductal gray. Distally, clusters of labeled cells were found ipsilaterally in the caudal periaqueductal gray extending to the nucleus cuneiformis, and bilaterally in the locus coeruleus and supramamillary decussation. More widespread labeling was found in most hypothalamic subareas and in the lateral habenula. The labeled brain areas following ventral tegmental area stimulations were totally distinct, and comprised the medial forebrain bundle, the nucleus accumbens, the vertical limb of the diagonal band and the medial septum. The pattern of labeling produced by periaqueductal gray stimulations was therefore specific, and provided information about brain structures involved in the motivational and behavioral effects of such stimulations.


Membrane-bound choline acetyltransferase of the torpedo has characteristics of an integral membrane protein and can be solubilized by proteolysis.

Due to Triton X-114 fractionation of synaptosomes isolated from the electric organ of the fish Torpedo, the existence of a hydrophilic and an amphiphilic form of the enzyme choline-O-acetyltransferase (ChAT) was revealed. Amphiphilic ChAT which represents about 10% of total enzyme activity in synaptosomes, reached 40% of ChAT activity measured in preparations of synaptosomal plasma membranes (SPM) which were washed with solutions of increasing ionic strength. ChAT activity bound to washed SPM could be partially solubilized using proteinase K but not phospholipase C. No ChAT solubilization occurred by treating intact synaptosomes with proteinase K. Water/Triton X-114 partition coefficients of hydrophilic and amphiphilic ChAT were found to be 6.5 and 0.17, respectively. Sedimentation coefficients determined by centrifugation in linear density gradients of sucrose containing Triton X-100, were 4.2S and 4.4S for amphiphilic and hydrophilic ChAT, respectively. On the other hand, removal of Triton X-114 from the detergent phase containing amphiphilic ChAT activity led to enzyme aggregation. Finally, amphiphilic ChAT was slightly more acidic (pH 6.6) than was hydrophilic enzyme (6.8-7.0). We conclude that in Torpedo synaptosomes two forms of ChAT activity, a soluble and a membrane-bound form, are indeed present which differ in their hydrophobicity. The soluble form is hydrophilic. The membrane-bound form is amphiphilic and it aggregates upon removal of detergent. These are two characteristics of integral membrane proteins. Membrane-bound ChAT is most probably intracellularly oriented and not bound to membrane through a 'receptor' protein.


Significance of an opiate mechanism in the adjustment of cerebrocortical oxygen consumption and blood flow during hypercapnic stress.

The role of adrenal medulla-derived enkephalins in the control of hypercapnic cerebrocortical blood flow (CBF) and oxygen consumption (CMRO2) was investigated in the ketamine anesthetized rat. Three experimental interventions were utilized: inhibition of opioid receptors with naloxone, decrease of adrenal enkephalin production with chronic adrenal medullectomy, and treatment of adrenal demedullated animals with the synthetic enkephalin analog, D-Ala2, N-Me-Phe4, Gly5-ol-enkephalin (DAGO). In intact, untreated animals hypercapnia increased CBF and CMRO2 by approximately 300 and 35%, respectively. Naloxone reduced the hypercapnic increase of CBF, and transformed the hypercapnic increase of CMRO2 into a decrease. The mid-points of the dose-response curves for (1)-naloxone and (d)-naloxone were 10 micrograms/kg and 100 micrograms/kg, respectively. Adrenal demedullation and treatment with (1)-naloxone (0.2 mg/kg) decreased the hypercapnic CBF and CMRO2 by approximately 50%. DAGO treatment of adrenal demedullated animals restored the hypercapnic CBF and CMRO2 to values similar to those found in intact animals. These observations suggest that opioid peptides (most likely adrenal medulla-derived enkephalins) play a significant role in the regulation of CMRO2 and CBF during moderate hypercapnia.


Electrophysiology of dentate granule cells after kainate-induced synaptic reorganization of the mossy fibers.

Morphological data from humans with temporal lobe epilepsy and from animal models of epilepsy suggest that seizure-induced damage to dentate hilar neurons causes granule cells to sprout new axon collaterals that innervate other granule cells. This aberrant projection has been suggested to be an anatomical substrate for epileptogenesis. This hypothesis was tested in the present study with intra- and extracellular recordings from granule cells in hippocampal slices removed from rats 1-4 months after kainate treatment. In this animal model, hippocampal cell loss leads to sprouting of mossy fiber axons from the granule cells into the inner molecular layer of the dentate gyrus. Unexpectedly, when slices with mossy fiber sprouting were examined in normal medium, extracellular stimulation of the hilus or perforant path evoked relatively normal responses. However, in the presence of the GABAA-receptor antagonist, bicuculline, low-intensity hilar stimulation evoked delayed bursts of action potentials in about one-quarter of the slices. In one-third of the bicuculline-treated slices with mossy fiber sprouting, spontaneous bursts of synchronous spikes were superimposed on slow negative field potentials. Slices from normal rats or kainate-treated rats without mossy fiber sprouting never showed delayed bursts to weak hilar stimulation or spontaneous bursts in bicuculline. These data suggest that new local excitatory circuits may be suppressed normally, and then emerge functionally when synaptic inhibition is blocked. Therefore, after repeated seizures and excitotoxic damage in the hippocampus, synaptic reorganization of the mossy fibers is consistently associated with normal responses; however, in some preparations, the mossy fibers may form functional recurrent excitatory connections, but synaptic inhibition appears to mask these potentially epileptogenic alterations.


TCP enhances the survival of human fetal spinal cord cells in culture.

Dissociated cell cultures were prepared from human spinal cords of 7-10-week-old fetuses. After 10 weeks progressive neuronal necrosis was observed in controls whereas N/1-(2-thienyl)cyclohexyl/piperidine (TCP) enhanced the survival time of the cells. After 21 weeks the number of gamma-aminobutyric acid (GABA)ergic and neuron specific enolase (NSE)-stained neurons was higher in the TCP-treated cultures than in controls. TCP appears to be a promising drug for long term survival of neurons.


Serotonergic phase shifts of the mammalian circadian clock: effects of tetrodotoxin and high Mg2+.

The mammalian circadian clock in the suprachiasmatic nuclei (SCN) can be phase-shifted in vitro by the serotonin agonist quipazine. Here we show that quipazine resets the SCN clock in the presence of tetrodotoxin or 10 mM Mg2+, treatments that block Na+ action potentials and Ca2+ channels, respectively. These results support the hypothesis that quipazine resets the clock by stimulating receptors located on clock elements rather than on cells afferent to the clock.


5 alpha-pregnane-3 alpha, 21-diol-20-one (THDOC) attenuates mild stress-induced increases in plasma corticosterone via a non-glucocorticoid mechanism: comparison with alprazolam.

5 alpha-Pregnane-3 alpha,21-diol-20-one (THDOC; 5 mg/kg) and the triazolobenzodiazepine alprazolam (1 mg/kg) attenuated mild stress-induced increases in plasma corticosterone concentrations via GABAergic mechanisms. Unlike alprazolam, THDOC failed to decrease corticotropin-releasing factor (CRF) concentrations in the locus ceruleus. While THDOC may plausibly act via endogenous GABAergic mechanisms to reduce stress-induced endocrine and behavioral responses that are likely mediated in part by CRF neurons, these preliminary findings suggest that, at the dose and time point studied, THDOC does not identically mimic the actions of alprazolam, another drug which potentiates GABAergic activity.


Horseradish peroxidase labeling of growth cones and axons beyond the site of injury in injured rabbit optic nerve axons growing in their own environment.

Spontaneous growth of injured axons in the mammalian central nervous system is limited. We have previously shown an apparently regenerative growth of injured optic axons in the adult rabbit, achieved by supplying them with soluble substances originating from growing axons, followed by low energy helium-neon laser irradiation. The growing unmyelinated and thinly myelinated axons were embedded in astrocytes, and some were in the process of remyelination by oligodendrocytes. They were shown to have originated from the retinal ganglion cells. The present study further supports evidence relating to the origin and nature of these axons. Light microscopic analysis of these axons labeled with anterogradely transported horseradish peroxidase revealed that many of these axons have varicosities and bear growth cone-like swellings in their tips. These axons traverse the lesion site and extend into the distal stump in a disorganized pattern.


Protection against dendrotoxin-induced clonic seizures in mice by anticonvulsant drugs.

Various anticonvulsant drugs were evaluated for their ability to protect against clonic seizures induced in mice by intraventricular injection of the K+ channel blocking peptide dendrotoxin (DTX). Phenytoin, the phenytoin-like anticonvulsant carbamazepine and the broad spectrum drug valproate were effective in this model, whereas the GABA-enhancers diazepam and tiagabine, the NMDA antagonists (+/-)-CPP and (+)-MK-801, the AMPA antagonist NBQX, the antiabsence drug ethosuximide and the Ca2+ channel antagonist nimodipine were inactive. In contrast to the lack of activity of other NMDA antagonists, phencyclidine and ADCI [(+/-)-aminocarbonyl-10,11-dihydro-5H-dibenzo [a,d]cyclohepten-5,10-imine] were potent antagonists of DTX-induced seizures.


Intracerebral transplants of primary muscle cells: a potential 'platform' for transgene expression in the brain.

After the transplantation of rat primary muscle cells into the caudate or cortex of recipient rats, the muscle cells were able to persist for at least 6 months. Muscle cells transfected with expression plasmids prior to transplantation were able to express reporter genes in the brains for at least 2 months. These results suggest that muscle cells might be a useful 'platform' for transgene expression in the brain.


Kindling of the massa intermedia of the thalamus in rats.

The kindling response of the massa intermedia (MI) was assessed in rats. Clinical manifestation of the MI kindling was generally similar to that of limbic kindling, and positive transfer to the amygdala (AM) was obtained following MI kindling. However, MI kindling showed (1) a relatively high after discharge threshold which sometimes increased during the course of kindling, (2) a seizure stage instability with frequent regression to earlier stages, (3) a failure to establish a generalized seizure triggering threshold with an 'all-or-none' property, and (4) a generalized tonic-clonic seizure, which was quite different from a kindled limbic seizure, during early phase of kindling. Furthermore, the MI stimulation caused violent beating movement of the forelimbs, jumping, or running regardless of presence/absence of afterdischarge. The findings suggest that mechanisms other than a simple activation of limbic structures are involved in the process of MI kindling.


Evidence that fine primary afferent axons innervate a wider territory in the superficial dorsal horn following peripheral axotomy.

Peripheral axotomy initiates changes in central primary afferent receiving areas of the dorsal horn of the spinal cord. Most of the presently known changes are degenerative in nature and consist of such things as cell and axon death or declines in peptides or enzymes. Other changes are regenerative in nature and because most of these occur in the superficial dorsal horn, which is where fine primary afferents end, we wished to ask whether peripheral axotomy results in a change in the distribution in these fine afferents. Using recently available markers for fine primary afferent axons and small dorsal root ganglion cells, we demonstrate that peripheral axotomy results in a considerable increase in the immunolabeled area for these compounds. Our interpretation is that there may be an extension of fine primary afferent fibers into lamina III and possibly lamina IV following peripheral axotomy. If further work bears out this conclusion, this would provide a possible explanation for the chronic pain states that sometimes follow peripheral nerve damage.


Preservation of hippocampal brain slices with in vivo or in vitro hypothermia.

Hippocampal brain slices show CA1 injury similar to that seen after global ischemia in vivo. Cooling rats to 31 degrees C prior to sacrifice or cooling slices to 21 degrees C for 45 min increased the percentage of normal CA1 pyramidal cells after 5 h in vitro from 30% to over 80%. Brain slices also show a unique, consistent injury in dentate which is lessened by transient cooling to 21 degrees C but not by cooling the animal.


Evidence for different sites of monoamine oxidase and catechol-O-methyltransferase in the striatum.

Wistar rats were implanted with a probe for brain microdialysis in the striatum, and measured for two major dopamine metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA). Intraperitoneal chlorpromazine (5 mg/kg) increased the concentrations of both metabolites in the dialysate when the determinations were carried out before 90 min after the administration. An apparent difference between the declines in concentrations of DOPAC and HVA was observed. Although the concentration of HVA maintained higher levels for over 6 h after the administration, DOPAC underwent a decrease in its concentration after the maximum level, which occurred at 90 min after the administration. These results are discussed in terms of the different distributions of two enzymes related to the metabolism of dopamine, catechol-O-methyltransferase and monoamine oxidase.


Different effects of hypoxia on the membrane potential and input resistance of isolated and clustered carotid body glomus cells.

Intracellular recordings were made from cultured glomus cells of rat carotid bodies. Hypoxia (PO2 1-61 Torr), induced by Na-dithionite (Na2S2O4), differentially affected cells that were clustered and those that were isolated. More than 80% of clustered cells depolarized and their input resistance decreased, whereas about 60% of isolated cells hyperpolarized and their input resistance increased. In both groups, the mechanisms for cell depolarization or hyperpolarization appeared similar. Differences between clustered and isolated cells seemed to be conditioned by the sustentacular cells. Their processes surrounded clustered cells but were absent around isolated ones. However, we may have also functionally different glomus cells which may or may not correspond to the different types described by anatomists.


Brown adipose tissue temperature responses following electrical stimulation of ventromedial hypothalamic and lateral preoptic areas or after norepinephrine infusion to Long Evans or Sprague-Dawley rats.

Thermoregulatory (brown adipose tissue temperature) experiments were conducted in age-matched anesthetized, male Sprague-Dawley and Long Evans rats kept at 37 degrees C given unilateral electrical stimulation of the ventromedial hypothalamic nucleus or the lateral preoptic area or after intravenous infusion of norepinephrine HCl (50 micrograms/kg total dose). Unilateral electrical stimulation (0.5 ms pulses of 250 microA at 50 Hz for 30 s) of the ventromedial nucleus or lateral preoptic area caused significant rises (greater than 0.6 degrees C) of intrascapular brown adipose tissue temperature in the Long Evans group only. Norepinephrine infusion (0.1 ml/min for 10 min), or intravenous bolus injection of propranolol HCl (2.5 mg/kg) caused significant yet similar increases (greater than 1.0 degrees C) and decreases (greater than -0.3 degrees C), respectively, of intrascapular brown adipose tissue temperature from pre-administration control temperatures in both Long Evans and Sprague-Dawley groups. The results demonstrate that intrascapular brown adipose tissue sensitivity towards exogenous norepinephrine or propranolol administration is similar between Sprague-Dawley and Long Evans rats acclimated to 21 degrees C as determined by increases in temperature in the intrascapular brown adipose tissue pad. However, intrascapular brown adipose tissue thermogenesis of 21 degrees C-acclimated Long Evans rats is also activated after CNS electrical stimulation of the ventromedial nucleus or lateral preoptic area, an activation not seen in Sprague-Dawley rats suggestive that the latter group has some inhibition of neural pathways that activate brown adipose tissue thermogenesis.


Acute and persistent effects of methamphetamine on developing monoaminergic neurons in reaggregate tissue culture.

Three-dimensional, rotation-mediated reaggregate tissue cultures composed of rostral mesencephalic cells and corpus striatal cells were used to examine the short-term and persistent effects of methamphetamine on developing monoamine-containing neurons. Reaggregates were exposed to drug for one week. Reductions in reaggregate endogenous dopamine and serotonin levels occurred following treatment with methamphetamine during days 15-22 of culture over the concentration range 10(-7) to 10(-4) M. The highest methamphetamine concentration reduced dopamine and serotonin levels to 29 and 33%, respectively, of control values. Monoamine levels were reduced from control values after 3 days of exposure to 10(-4) M methamphetamine. No further reduction resulted from 4 additional days of drug treatment. In order to determine whether monoaminergic neurons would recover from the drug-induced deficit, reaggregates were exposed to 10(-4) M methamphetamine for 7 days and then grown in drug-free media for an additional 20 days. During the 20 day recovery period, monoamine levels in the control group increased with time in culture. After an initial rapid increase (recovery days 0-9), the level of monoamines in the recovery group remained at a constant proportion to the level in the control group suggesting that the monoaminergic neurons return to a rate of development similar to that seen in untreated cultures. However, this rate was not sufficient to overcome the reduction in monoamine levels produced by 7 days of methamphetamine treatment. The results indicate that the effects of methamphetamine on developing monoaminergic neurons are marked and persistent.


Corticotropin-releasing factor mediated muscle atonia in pons and medulla.

The dorsolateral pontine inhibitory area (PIA) and medial medullary reticular formation (MMRF) have been found to mediate the muscle atonia of REM sleep. Our previous studies have shown that acetylcholine (ACh) microinjection in the PIA and in the nucleus paramedianus of the medial medulla produces muscle atonia. Glutamate microinjection in both PIA and nucleus magnocellularis (NMC) of the medial medulla also produces muscle atonia. Since immunohistochemical studies have identified corticotropin-releasing factor (CRF) as a potential dorsolateral pontine and NMC transmitter, the present study was undertaken to determine whether this transmitter could produce suppression of muscle tone. Experiments were performed on unanesthetized, decerebrated cats. CRF was microinjected into points in the PIA and NMC at which electrical stimulation produced bilateral inhibition of muscle tone. We found that CRF produced a dose-dependent muscle tone suppression. At 10 nM concentration, the latency and duration of muscle inhibition produced by CRF injection were comparable with those of L-glutamate, at 18.8 s and 4.1 min, respectively. This CRF-induced muscle inhibition was blocked by the CRF antagonist, alpha-helical [Glu27]corticotropin-releasing factor 9-41 (CRF 9-41). Microinjection of CRF and non-NMDA agonists, kainate and quisqualate, into the same sites in PIA and NMC produced muscle atonia. Pontine sites at which CRF injection induces atonia are identical to those at which acetylcholine microinjection produces atonia. These results indicate that CRF may interact with glutamate and acetylcholine in the generation of muscle atonia.


Acute and prolonged effects of ibogaine on brain dopamine metabolism and morphine-induced locomotor activity in rats.

Ibogaine, an indolalkylamine, proposed for use in treating opiate and stimulant addiction, has been shown to modulate the dopaminergic system acutely and one day later. In the present study we sought to systematically determine the effects of ibogaine on the levels of dopamine (DA) and the dopamine metabolites 3,4 dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in tissue at several time points, between 1 h and 1 month post-injection. One hour after ibogaine-administration (40 mg/kg i.p.) a 50% decrease in DA along with a 37-100% increase in HVA were observed in all 3 brain regions studied: striatum, nucleus accumbens and prefrontal cortex. Nineteen hours after ibogaine-administration a decrease in DOPAC was seen in the nucleus accumbens and in the striatum. A week after administration of ibogaine striatal DOPAC levels were still reduced. A month after ibogaine injection there were no significant neurochemical changes in any region. We also investigated the effects of ibogaine pretreatment on morphine-induced locomotor activity, which is thought to depend on DA release. Using photocell activity cages we found that ibogaine pretreatment decreased the stimulatory motor effects induced by a wide range of morphine doses (0.5-20 mg/kg, i.p.) administered 19 h later; a similar effect was observed when morphine (5 mg/kg) was administered a week after ibogaine pretreatment. No significant changes in morphine-induced locomotion were seen a month after ibogaine pretreatment. The present findings indicate that ibogaine produces both acute and delayed effects on the tissue content of DA and its metabolites, and these changes coincide with a sustained depression of morphine-induced locomotor activity.


5-HT2 receptors in the nucleus tractus solitarius: characterisation and role in cardiovascular regulation in the rat.

The effects of the local application of drugs acting on 5-HT2 receptors in the nucleus tractus solitarius (NTS) on the heart rate and blood pressure were investigated in normal and nodose ganglionectomized anaesthetized rats. The unilateral micro-injection of an agonist such as 2,5-dimethoxy-3-bromo-amphetamine (DOB) (0.1-0.5 pmol) or 2,5-dimethoxy-3-nitroamphetamine (DON) (0.1-0.5 pmol) produced a dose-dependent hypotension and bradycardia in both intact and ganglionectomized animals. These cardiovascular effects were similar to those observed after the unilateral micro-injection of low doses (pmol) of 5-HT, and could be prevented by the prior micro-injections of the 5-HT2 antagonists ketanserin, ritanserin and piremperone. These findings support the hypothesis that 5-HT2 receptors within the NTS play a role in the reflex regulation of blood pressure. In addition, it was also observed that the micro-injection of subthreshold doses of 5-HT or DOB significantly enhanced the hypotension and bradycardia produced by the unilateral micro-injection of N-methyl-D-aspartate (NMDA). The potentiation of NMDA depressor effects by 5-HT or DOB could be totally prevented by ketanserin or piremperone, suggesting that 5-HT acting upon 5-HT2 receptors in the NTS may intervene in the reflex control of blood pressure by modulating the glutamatergic transmission.


Is CNS trauma a prerequisite for the elongation of CNS axons into denervated peripheral nerve?

The demonstration that some central nervous system (CNS) axons can regenerate when provided with a suitable environment raises the possibility of new treatments for CNS injury. However, at present the conditions for optimal regeneration are not well understood. For example, the methods used in previous studies have entailed CNS trauma as part of the research protocol (e.g. that resulting from the implantation of peripheral nerve grafts), and so the role of neuronal or axonal injury in the regrowth observed has been difficult to establish. To determine whether such injury is necessary for the central reinnervation of denervated peripheral nerve, the L5 dorsal root has been chronically denervated in rats by freeze-thawing its dorsal root ganglion (DRG), and the root has been left attached to either traumatized or non-traumatized spinal cord. The trauma induced was quite mild, and resulted from several vertical insertions of a fine needle. Two to 4 months later, retrogradely transported horseradish peroxidase (HRP) was used to label spinal neurons which sent axons into the denervated roots. HRP-labelled neurons were found in each of the spinal cords subjected to trauma, but no labelled neurons were observed in any of the non-traumatized cords. The number of HRP-labelled neurons in individual spinal cords was positively correlated with the degree of spinal cord trauma. We conclude first that the chronic and intimate presence of a denervated PNS tissue in continuity with the spinal cord is not, in itself, a sufficient stimulus to induce its reinnervation by CNS axons. Second, we conclude that under the conditions of this experiment CNS trauma is a prerequisite for the reinnervation of denervated peripheral nervous tissue by CNS axons.


Differential sites of origin and collateralization of corticospinal neurons in the rat: a multiple fluorescent retrograde tracer study.

Cells of origin for corticospinal fibers in the rat were identified following retrograde transport of Fluoro-Gold (FG), Propidium iodide (PI), Fast blue (FB), and Diamidino yellow (DY) injected unilaterally into lumbar (FG), mid-thoracic (PI), cervical enlargement (FB), and cranial cervical (DY) spinal gray matter. Most labeled neurons were contralateral to injection in lamina V and ranged from small to very large. These cells occupied two distinct cortical regions: one rostral and the other larger and more caudal. Neurons of the rostral region projected axons solely to cervical spinal segments whereas neurons of the caudal region projected fibers to all spinal segments. Somatotopically, most neurons projecting to lumbar segments were most medial. More than 98% of all labeled cortical neurons contained only a single fluorescent tracer; however, within a single tissue section each of the 4 tracers could be found in these single labeled neurons. The few double labeled neurons contained only cervical (DY + FB) or thoracolumbar (PI + FG) tracers. No triple or quadruple labeled cells were seen. Hence morphological evidence is presented that corticospinal axons branch to terminate in more than one spinal region, but these collateral terminations are restricted to only a few adjacent spinal segments.


Distribution of cannabinoid receptors in rat brain determined with aminoalkylindoles.

Extensive mapping of the cannabinoid receptor in rat brain has been reported recently using synthetic cannabinoids. Another class of compounds, the aminoalkylindoles (AAIs), does not resemble the cannabinoids structurally. Ligand binding data on isolated membranes, however, indicate that AAIs bind to the cannabinoid receptor. The present experiments compared the binding of AAIs and synthetic cannabinoids in vitro and by receptor autoradiography. The AAIs bound to a receptor in rat cerebellum with high affinity (Kd = 15 nM), and synthetic cannabinoids were potent competitors for AAI binding sites. In the autoradiographic studies in rat brain, an AAI and a synthetic cannabinoid were used to compete for the binding of a radiolabeled AAI to compare regionally and quantitatively the inhibition of AAI binding by the two classes of compounds. The distribution of the AAI binding was very similar to that reported for synthetic cannabinoid binding. These data add further evidence that the aminoalkylindoles bind to the cannabinoid receptor. Furthermore, the autoradiographic data for AAI binding, in addition to the autoradiographic data for the synthetic cannabinoid, provide a high degree of confidence in the localization of the cannabinoid receptor in the rat brain.


Amphiphilic behavior of a brain tetrameric acetylcholinesterase form lacking the plasma membrane anchoring domain.

We have studied the behavior of a mammalian brain tetrameric acetylcholinesterase (AChE) form released by proteinase K from a crude membrane fraction of bovine caudate nucleus. The solubilization of active AChE indicated the presence of a protease-sensitive site in the anchored protein. Unexpectedly, the solubilized AChE maintained its capacity to form aggregates in detergent-free gradients. We demonstrate here that this property was due neither to the presence of the hydrophobic membrane-anchoring domain still linked to the enzyme, nor to the presence of AChE activity trapped in small plasma membrane vesicles. Moreover, we found that the proteinase K-treated extract, devoid of AChE activity, induced the aggregation of purified hydrophilic AChE which usually does not form aggregates. Our results suggest the presence of an AChE aggregating factor in bovine brain extracts prepared in the presence of proteinase K. It is possible that this aggregation may reflect a process of AChE clustering on neurons.


Infusions of 6-hydroxydopamine into the nucleus accumbens abolish the analgesic effect of amphetamine but not of morphine in the formalin test.

The effects of bilateral infusions of 6-hydroxydopamine into the nucleus accumbens on analgesia produced by D-amphetamine and morphine were examined, in separate experiments, in the formalin test in rats. The lesions in the two experiments were not significantly different and dopamine in the nucleus accumbens, olfactory tubercle and striatum was depleted to 21.1%, 40.3% and 65.0% of control values, respectively. D-Amphetamine (0.75 and 2.0 mg/kg) and morphine (3.0 and 6.0 mg/kg) attenuated the response to formalin in unlesioned control rats. The analgesic effect of amphetamine was severely reduced by 6-hydroxydopamine lesions, and the residual analgesia was correlated with the amount of dopamine in the nucleus accumbens, but not with dopamine levels in the olfactory tubercle or striatum. Lesions also attenuated the locomotor stimulant effect of amphetamine. The analgesic effect of morphine was not altered by 6-hydroxydopamine infusions, nor was there any correlation between the analgesic effect of morphine and dopamine concentration in the nucleus accumbens, olfactory tubercle or striatum. The results indicate that the dopamine innervation of the nucleus accumbens is not critical for the analgesic effect of morphine but plays a major role in the analgesic effect of amphetamine.


Insulin-like peptide(s) in the central nervous system of the snail Helisoma duryi.

The central nervous system of the snail Helisoma duryi contains porcine insulin-immunoreactive cells. Most of these cells are known as neurosecretory mediodorsal cells (MDC), and are involved in regulation of growth. There are about 25-40 large and small MDC in each cerebral ganglion near the commissure. Besides the MDC, 6-8 insulin-immunoreactive cells are also seen in the left parietal ganglion and the visceral ganglion. Insulin-immunoreactive material can be traced from the MDC perikarya into their axons in the median lip nerve. The elementary granules of the MDC are 150-250 nm in diameter, and are insulin-immunoreactive. Insulin-like material is released by the CNS in vitro, and can be measured by radioimmunoassay using an antiserum against human insulin. High potassium and 4-aminopyridine increase such release in vitro. It is likely that insulin-like peptide is a growth hormone in H. duryi.


Chronic neonatal MK-801 treatment results in an impairment of spatial learning in the adult rat.

Chronic neonatal treatment with the non-competitive N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 from postnatal day 8 through 19 has been shown to affect hippocampal NMDA receptor function of adult rats. Since many studies have shown that NMDA receptors play a crucial role in learning and memory, and since one of the hippocampal functions is spatial learning, we have examined whether this changed response of hippocampal neurons is associated with changes in its normal function. We therefore tested spatial learning and memory using a water maze in adult rats neonatally treated with MK-801. MK-801-treated rats were able to learn the spatial task as well as control rats but at a significantly slower rate. Performance in a visual cue task was not affected by the neonatal treatment, suggesting that the slower spatial learning is not caused by locomotor or sensory deficits. These results suggest that chronic NMDA receptor blockade during the neonatal period leads to long-lasting disturbances of hippocampal function.


The effects of A- and C-fiber stimulation on patterns of neuropeptide immunostaining in the rat superficial dorsal horn.

The present study determines the effects of sciatic nerve stimulation at intensities that activate A-fibers alone or both A- and C-fibers on immunostaining for substance P (SP), cholecystokinin-octapeptide (CCK-8), galanin (GAL), dynorphin (DYN) and vasoactive intestinal polypeptide (VIP) in the superficial dorsal horn of the rat spinal cord. The goal of this study is to provide a more precise spatial localization of the sites of release or accumulation of these compounds in relation to specific types of stimuli. Following A-fiber stimulation, there was no significant change in immunostaining for any of these compounds. However, A- and C-fiber stimulation resulted in major changes. For SP, CCK-8, GAL and DYN there was a large and significant loss of immunostaining in medial regions of the dorsal horn. This is the area where sciatic nerve primary afferent fibers terminate and the depletion is probably correlated with activity in these fibers. By contrast, VIP immunostaining is increased in the lateral part of the superficial cord, which is outside of the central sciatic afferent fiber terminations. This indicates that the increase is not in the fine sciatic sensory axons that are directly stimulated. As a final point, the fact that C-fiber but not A-fiber stimulation causes marked changes in the immunocytochemical distribution of all these compounds is further evidence, albeit indirect, that they are involved in nociceptive information processing.


Electrophysiological and anatomical studies on thalamic mediodorsal nucleus projections onto the prefrontal cortex in the cat.

Electrical stimulation of the mediodorsal nucleus (MD) of the thalamus elicited field potentials in the gyrus proreus (PRO), frontalis (FR), rectus (RE) and cinguli anterior (CIant) of the ipsilateral prefrontal and adjacent cortical areas in cats. The results of a laminar field potential analysis indicate that the field potentials can be regarded as a combination of deep and superficial thalamocortical responses. By injecting horseradish peroxidase (HRP) into the MD, HRP-labeled terminals were distributed in the prefrontal and adjacent cortical areas where the field potentials were elicited. Densely labeled terminals in cortical layer I were distributed where the superficial thalamocortical responses were prominent, while those in layers III-V were distributed in the areas where the deep thalamocortical responses were prominent.


Correlation between the induction of an immediate early gene, zif/268, and long-term potentiation in the dentate gyrus.

Expression of the immediate early gene zif/268 (also termed NGFI-A, Krox 24, TIS8 and Egr-1) was investigated in awake rats following various long-term potentiation (LTP) induction protocols. zif/268 mRNA (Northern blots) and protein (immunohistochemistry) levels sharply increased following LTP, and followed a time course characteristic of other immediate early genes. When measured across 3 tetanization protocols known to produce differing degrees of LTP persistence, zif/268 induction was found to be more highly correlated with LTP duration than with the magnitude of initial LTP. These data support the hypothesis that the immediate early gene zif/268 plays a role as a third messenger in the cascade of cellular and nuclear events that govern the persistence of LTP.


Ibotenic acid lesions in the amygdaloid central nucleus but not in the lateral subthalamic area prevent the acquisition of differential Pavlovian conditioning of bradycardia in rabbits.

The present study examined the effect of ibotenic acid lesions in the amygdaloid central nucleus (ACe) or in the lateral zona incerta of the subthalamus (LZI) on the acquisition of differential Pavlovian conditioning of bradycardia in rabbits. Previous work has shown that bilateral electrolytic lesions in either ACe or LZI abolished the retention of conditioned heart rate (HR) responses. In order to determine whether these findings were due to destruction of cells intrinsic to ACe or LZI, ibotenic acid lesions were placed bilaterally in either structure or in control sites. Following recovery, animals were subjected to differential Pavlovian conditioning in which one tone (CS+) was paired with periorbital shock and a second tone (CS-) was presented alone. It was found that destruction of cell bodies in ACe, but not LZI, prevented the acquisition of the differential bradycardiac conditioned response. In addition, ACe lesions did not interfere with baseline HR, the HR orienting response, the HR unconditioned response to shock, or the concomitantly conditioned corneoretinal potential. The results of this study suggest that destruction of cells intrinsic to ACe selectively prevents the acquisition of differentially conditioned HR, and perhaps other conditioned responses related to conditioned arousal, but does not affect unlearned HR responses or specific somatomotor conditioned responses.


Male rat copulation following 6-OHDA lesions of the medial preoptic area: resistance to repeated administration and rapid behavioral recovery.

Dopamine (DA) in the medial preoptic area (MPOA) has been shown to facilitate male rat sexual behavior. However, injections of the catecholamine (CA) neurotoxin 6-OHDA into the MPOA did not impair copulation in tests 3 days after injection. In the present study, three weekly (serial) injections produced no copulatory deficits compared to animals that received a single injection or to preinjection copulatory behavior scores. However, blocking CA synthesis, which did not impair control rats, produced deficits in both single and serial lesion animals, with significantly fewer serial than single lesion animals initiating copulation. Biochemical analysis of tissue punches showed no difference in MPOA concentrations of dopamine, norepinephrine, epinephrine, or the dopamine metabolite DOPAC between the two groups. Additional animals were tested at earlier intervals after 6-OHDA injections into the MPOA. Tests conducted 30 min after an MPOA injection of 6-OHDA revealed that all measures of copulation were impaired, relative to scores 24 h later. However, these scores were not significantly different from animals tested 30 min after a vehicle injection. A final group, tested 4 h after injection, showed impairment of all measures of copulation compared to vehicle injections and to tests 24 h later. Furthermore, in the tests 24 h later, 6-OHDA animals were not different from vehicle animals. Results from all experiments show that 6-OHDA injections into the MPOA impair copulation for at least 4 h, but that behavioral recovery is complete 24 h later. However, deficits can be reinstated by inhibiting DA synthesis, suggesting that increased synthesis in undamaged terminals contributed to behavioral recovery.


The effect of stimulus duration on noxious-stimulus induced c-fos expression in the rodent spinal cord.

C-fos is a proto-oncogene that is expressed within some neurons following depolarization. The protein product, fos, has been proposed as an anatomical marker for neuronal activity following noxious peripheral stimulation. However, the literature on noxious-stimulus induced fos expression contains several puzzling observations on the time course and laminar distribution of neuronal labeling within the spinal cord. This study has analyzed the effect of stimulus duration on the expression of fos-like immunoreactivity (FLI) within the spinal cord of anesthetized rats. In order to examine the time course of fos expression following brief periods of stimulation, we required a type of stimulus that was intense enough to activate nociceptors but that did not produce tissue damage. We have therefore employed pulsed, high intensity electrical stimulation, with stimulus durations ranging from 3 s to 24 h. The results indicate that stimulus duration has a profound effect upon the number of labeled cells, the intensity of neuronal labeling, the laminar pattern of FLI, and the time course of fos expression. Brief stimulation periods induce relatively few and relatively lightly labeled neurons, located predominantly within the most superficial laminae of the dorsal horn. Maximal immunoreactivity appears approximately 2 h after stimulation has ceased, and disappears within hours. Continuous stimulation produces many more labeled cells, darker labeling, and FLI within both dorsal and ventral laminar regions. Maximal FLI is seen after approximately 4.5 h of continuous stimulation, with reduction in the number of labeled cells thereafter. These data indicate that the results of any study employing c-fos as a marker for neuronal activity may be affected by the duration of the exciting stimulus.


Impulses of variable amplitude recorded from cell-attached patches on small hippocampal neurons.

Small cultured neurons from embryonic rat hippocampus were investigated with the cell-attached configuration of the patch-clamp technique. The currents associated with spontaneous cellular impulse generation were studied. The positive peak current amplitudes varied by a factor larger than 2.5 for 11 of 12 cells quantitatively investigated, suggesting underlying action potentials of variable amplitude. A simple RC-network was found to describe the properties of the patch membrane, and was used to calculate the underlying potential time course. The action potentials computed showed a considerable variation in amplitude. The results suggest that the impulse variability previously analysed with the whole-cell technique was not caused by dialysis of the cell interior.


Myoclonus in the decerebrate cat produced by gallamine.

After intravenous infusion maintaining a neuromuscular blocking concentration very little gallamine enters the cerebrospinal fluid (CSF) of the intact anesthetized cat even after several days. After a similar intravenous infusion in the decerebrate cat gallamine enters the CSF slowly over 4 days eventually reaching a concentration similar to that in the plasma. This procedure is accompanied by very strong twitching in many muscles and the occurrence of slow waves in the inferior olive in synchrony with the twitches. A large dose (4 mg) of gallamine triethiodide injected directly into the cisterna magna of an intact anesthetized cat produced twitching within 1 min and slow waves in the inferior olive in good synchrony with the twitches; the effects lasted at least 43 h. Injection of a quantity of gallamine triethiodide (about 130 micrograms) sufficient to mimic the concentration in the CSF obtained after 3-4 days of neuromuscular block in the decerebrate cat (50-120 micrograms/ml gallamine) had extremely weak effects lasting for at most 1 h. However, this weak effect is probably due to the anesthetic because injection of a similar quantity into an (unanesthetized) decerebrate cat at any time after decerebration had strong effects. After intracisternal injection of gallamine the concentration in the CSF reaches very low levels within 12 h but twitching and activity in the inferior olive persists for 1-2 days. The reasons for this prolonged action are now being investigated. The effects of gallamine are compared with the condition of reticular reflex myoclonus.


Protective effects of brain hypothermia on behavior and histopathology following global cerebral ischemia in rats.

The present experiments were designed to assess whether brain hypothermia can reduce the behavioral and histopathological deficits associated with global forebrain ischemia. Animals were subjected to 12.5 min of four vessel occlusion (4VO) with moderate hypotension, and brain temperature maintained at either 37 degrees C (4VO-37) or 30 degrees C (4VO-30). Behavioral tests designed to assess forelimb reflexes and sensorimotor function were given on post-operative weeks 2 and 4. Beginning in week 5, the rats were trained on a variety of navigation problems in the Morris water maze. Histopathological examination of the tissue 2 months following reperfusion revealed that 4VO-37 animals sustained substantial cell death in hippocampal region CA1 and moderate damage to the dorsolateral neostriatum. 4VO-30 animals showed minimal cell death in CA1 and neostriatum. There were no group differences for any of the sensorimotor measures, or for acquisition performance on either the simple place task or visible platform version of the water maze. In contrast, during acquisition of the learning set task, the performance of 4VO-37 animals was impaired relative to either of the other groups, whereas the performance of 4VO-30 animals was not significantly different from the sham controls. These data suggest that moderate intra-ischemic brain hypothermia provides long-lasting protection from behavioral deficits as well as neuronal injury following transient global ischemia.


Cortical input to parvalbumin-immunoreactive neurones in the putamen of the squirrel monkey.

The cortex projects heavily to the striatum and makes asymmetrical synaptic contact mainly with the spines of medium-sized densely spiny neurones. The possibility exists that corticostriatal terminals also make synaptic contact with classes of striatal interneurones. The primary objective of the present experiment was to determine whether parvalbumin-immunoreactive neurones, which represent a class of GABAergic interneurones in the striatum, also receive a direct synaptic input from corticostriatal fibres. The anterograde tracer biocytin was injected into the motor and premotor cortices of the squirrel monkey (Saimiri sciureus). Following perfuse-fixation, sections of the striatum were processed histochemically to reveal the transported biocytin using an avidin-biotin-peroxidase complex and diaminobenzidine as the chromogen. They were then immunostained to reveal parvalbumin using benzidine dihydrochloride as the chromogen. In both the light and electron microscopes, the morphological features and the afferent synaptic input of the parvalbumin-immunoreactive neurones were similar to those observed in other species. Similarly, the morphology and postsynaptic targets of the corticostriatal terminals were similar to those described in other species. Light microscopic examination revealed that the anterogradely labelled corticostriatal terminals were often in close apposition to the parvalbumin-positive neurones. At the electron microscopic level the biocytin-positive corticostriatal terminals were found to make asymmetrical synaptic contacts mainly with spines. The parvalbumin-positive neurones were seen to have an invaginated nucleus, extensive cytoplasm and relatively few spines. Parvalbumin-immunoreactive dendrites received a dense synaptic input consisting mainly of asymmetric synapses and only a few symmetric synapses. Biocytin-labelled corticostriatal terminals were often seen in asymmetrical synaptic contact with parvalbumin-immunoreactive dendrites. These results show that GABAergic interneurones identified on the basis of parvalbumin immunoreactivity, in addition to the projection neurones of the striatum, are under the direct influence of the cerebral cortex.


A novel monoclonal antibody against carbohydrates of L1 cell adhesion molecule causes an influx of calcium in cultured cortical neurons.

We have studied the function of carbohydrates of the L1 molecule, a member of the immunoglobulin superfamily of adhesion molecules, using a novel monoclonal antibody, mAb-L1(2E12), against L1 molecule. This antibody was specific for the 200 kDa component of mouse L1 molecule and its epitope was N-linked for complex-type oligosaccharides. The mAb-L1(2E12) was found to induce a rise in intracellular Ca2+ concentration ([Ca2+]i) in cultured mouse embryonic cortical neurons. The rise in [Ca2+]i was dependent on the concentrations of mAb-L1(2E12). The rise seemed to be due to an influx of extracellular Ca2+ as EGTA treatment abolished it. Both cadmium and nifedipine blocked the effect of mAb-L1(2E12), suggesting the Ca2+ influx was through voltage-operated Ca2+ channels, particularly L-type Ca2+ channels. These results provide an important insight for understanding the mechanisms by which oligosaccharides of the L1 molecule influence various functions of neural cells.


Neural and endocrine sensitivities to opioids decline as a function of multiparity in the rat.

Hormonal changes during pregnancy regulate the onset of maternal behavior at parturition. In addition, the concentrations of beta-endorphin and mu opioid receptors are higher during pregnancy and lower during lactation. Previous studies have shown that sensitivity of female rats to the disruptive behavioral effects of morphine changes as a function of the number of pregnancies and/or lactations the females undergo. The objectives of the present study were to determine whether central infusions of the endogenous opioid, beta-endorphin, would disrupt maternal behavior. Next, we investigated the possibility that the neural sensitivity to beta-endorphin changes with repeated pregnancies. And finally, we examined whether opioid-mediated endocrine responses also change as a function of multiparity. In the first study, bilateral infusions of low doses (0.06-0.72 nmol) of beta-endorphin into the medial preoptic area (MPOA) of lactating, primiparous rats disrupted maternal behavior. When comparable doses of beta-endorphin were infused into the MPOA of age-matched, multiparous rats, the behavioral effects of beta-endorphin were significantly attenuated. In response to suckling stimulation, an opioid-mediated endocrine response, primiparous mothers secreted more prolactin than did multiparous rats. Moreover, multiparous, but not primiparous, mothers were insensitive to the ability of naloxone, an opiate antagonist, to block suckling-induced increases in prolactin. These findings indicate that reductions in neural sensitivity to opioids develop as females undergo repeated pregnancies and lactations, changes which affect both behavioral and endocrine functions.


The functional anatomy of manual motor behavior after unilateral frontal lobe lesions.

In normal brains different behavioral tasks lead to increased metabolism in different loci. Resting monkeys with acute hemineglect following periarcuate lesions show deficient glucose utilization (LCGU) in motor thalamus and other loci of the hemisphere with the lesion. To determine if the abnormal LCGU could be altered by a specific behavioral task, we used behavioral testing and training, and quantitative 2-deoxyglucose (2-DG) autoradiography. Animals reached criterion in a bimanual traction task (BMTT), then were tested for hand preference and responses to sensory stimuli, and received unilateral frontal or sham lesions. Operated animals showed hemineglect, and, on BMTT, intermanual coordination errors. 2-DG of frontals at rest showed deficient LCGU in 4 of 7 thalamic, and 1 of 4 cortical regions assessed. In contrast, only 2 thalamic and no cortical regions were deficient in pulling animals. Thus, in the face of structural damage and distant deficient neural activation, performance of a specific behavioral task may increase metabolism in specific brain loci, implying that plasticity of neural activation vis-a-vis behavioral demand is present even acutely after brain injury.


Evoked responses from an in vitro slice preparation of a primary gustatory nucleus: the vagal lobe of goldfish.

The vagal lobe of goldfish (Carassius auratus) is a laminated structure in which primary gustatory afferents terminate in a stereotypical pattern. Because the afferent fibers enter and distribute within the lobe in the transverse plane, the structure appeared suitable for in vitro slice electrophysiology. Slices were cut on a vibratome at 400-800 microns thickness and placed in a fresh water teleost Ringer's solution. Following a recovery period, clear population responses were recorded following electrical stimulation of the incoming fiber bundle. The later two components of this evoked waveform were eliminated by removal of the calcium from the bathing solution indicating the synaptic origin of these potentials. Further, the waveform was highly dependent on the position of the recording electrode, both in terms of laminar and tangential position. Evoked response maxima corresponded to the layers in which the primary afferent fibers terminate. In addition, the maximal evoked response was limited to a tangential distance of approximately 100 microns. The spatial restriction of the evoked waveform therefore corresponds well with the known anatomical organization of the primary gustatory afferent fibers. The evoked waveforms are sensitive to stimulus repetition rate, being facilitated by stimulus trains less than 50 ms in duration and fatigued by stimulus repetitions as slow as 1 Hz. This in vitro preparation should provide a means for investigating the physiological and pharmacological properties of primary gustatory fibers and nuclei.


Morphological and functional development of the suprachiasmatic nucleus in transplanted fetal hypothalamus.

The development of the suprachiasmatic nucleus (SCN) in fetal rat hypothalamus transplanted to the adult brain was studied using morphological and functional methods. Anterior hypothalamic tissue was transplanted into the third ventricle, lateral ventricle or subarachnoid space of intact, adult hosts from E17 fetuses. These transplants developed the cytoarchitectonic and immunohistochemical staining characteristics of SCN, clusters of parvocellular neurons expressing vasopressin- and vasoactive intestinal polypeptide-like immunoreactivity in adjacent cellular populations, irrespective of the exact location of the transplanted tissue in the host brain. The functional status of the transplants placed in the rostral third ventricle and the foramen of Monroe was analyzed and compared to host SCN using in vitro recording of neuronal firing rate and measurement of metabolism using the 2-deoxyglucose (2-DG) technique. During subjective day, neuronal firing rates and 2-DG uptake were high in discrete cell groups within the transplants which were subsequently demonstrated to exhibit the cytoarchitectonic and immunohistochemical characteristics of SCN. The firing rates and 2-DG uptake in these areas were lower during the subjective night. This pattern of activity closely resembles that of the intact SCN. In contrast, neither transplanted anterior hypothalamic area, lacking an identifiable SCN-like structure, nor posterior hypothalamic area showed day-night differences in firing rate or 2-DG uptake. These observations indicate that SCN transplanted into intact adult hosts exhibits morphological and functional differentiation nearly identical to the host and that the transplanted SCN maintains circadian function which is probably entrained to the host SCN.


Plasminogen activators in the neuromuscular system of the wobbler mutant mouse.

Wobbler, the neurological mutant mouse, carries an autosomal recessive gene (wr) and has been characterized as a model of lower motoneuron disorders with associated muscle atrophy, denervation and reinnervation. During normal murine neuromuscular development a decrease in muscle plasminogen activator (PA) activity accompanies synapse maturation. In contrast, experimental denervation in adult mice leads to an increase in muscle PA activity. The purpose of the present study was to determine the possible involvement of PAs in the denervation/reinnervation phenomena and motoneuron degeneration that characterize the wobbler mutant mouse. We determined the degree of innervation and its characteristics in wobbler mice by measuring choline acetyltransferase (ChAT) activity. We measured ChAT in the spinal cord as well as in two different muscles known to be differentially affected, biceps brachii and gastrocnemius. We found a sharp decrease of ChAT activity in both muscles but not in spinal cord extracts. We estimated the extent of sprouting by the silver/cholinesterase stain. Motoneuron terminal sprouting, not detected in normal animals, was present in 40% of the neuromuscular junctions in wobbler mice. We estimated specific PA activities in biceps brachii and gastrocnemius muscle extracts, as well as spinal cord extracts, using both an amidolytic assay and fibrin zymography. Increased PA, predominantly urokinase-PA (uPA), was observed in wobbler mouse muscle. A greater uPA was detected in biceps brachii muscle than in gastrocnemius muscle, which is less impaired by the mutation. There was no change in spinal cord PA, although tissue type PA (tPA) is the predominant PA type there.(ABSTRACT TRUNCATED AT 250 WORDS)


Modulation of net outward current in cultured neurons by angiotensin II: involvement of AT1 and AT2 receptors.

In this study we have used whole-cell, voltage-clamp procedures to determine the effects of angiotensin II (AII) on net outward current (I(no)) in neurons co-cultured from the hypothalamus and brainstem of 1-day-old rats. Ino is the sum of all inward and outward membrane currents (minus Na+, which is blocked by tetrodotoxin) which occur during the repolarization phase of the action potential. We have determined that AII elicits two separate effects on I(no) in cultured neurons. AII caused a reversible and concentration (0.1 nM-10 microM)-dependent increase in I(no). This effect is inhibited by the AT2 receptor-selective antagonists, PD123177 and PD123319 (both 100 nM), but not by the AT1-selective receptor blocker, DuP753 (Losartan; 100 nM), and so it is mediated by AT2 receptors. In a smaller number of neurons AII induced a reversible and concentration (0.01 nM-10 microM)-dependent decrease in I(no) that was blocked by Losartan (100 nM) but not by PD123177 (100 nM). Thus the decrease in I(no) is mediated by AT1 receptors. Additionally, some neurons displayed both AT1- and AT2 receptor-mediated effects on I(no). Our results demonstrate two distinct actions of AII on membrane ionic currents in cultured neurons, effects that are mediated by different AII receptor subtypes.


The effects of ibotenate lesions of the median preoptic nucleus on experimentally-induced and circadian drinking behavior in rats.

Male Sprague-Dawley rats were injected with either ibotenic acid or vehicle in the region of the median preoptic nucleus (MnPO) during methoxyflurane anesthesia. Later, the rats were tested for drinking responses elicited by angiotensin II (1 and 2 mg/kg s.c.) and hypertonic saline (3 and 6% w/v s.c.). Tests were conducted at 3 different phases of a 12:12 light-dark (LD) cycle (in the middle of the light phase and in the early and midportion of the dark phase). The rats with ibotenate lesions of the MnPO drank significantly less than vehicle-injected and lesion control groups regardless of when the tests were conducted. Subsequent monitoring of the diurnal rhythm of drinking, employing electrical lickometers, detected no difference between the rhythms of 4 rats with ibotenate lesions of the MnPO who failed to drink to homeostatic challenges and 4 vehicle-injected control rats. The results indicate that ibotenic acid lesions of the MnPO block drinking behavior stimulated by angiotensin II and hypertonic saline without disrupting the entrainment or pattern of ad libitum drinking.


Effect of chronic pre- and post-natal low-dose ethanol exposure on brain enolase isoenzyme activities.

Sprague-Dawley dams were treated with 3 v/v % ethanol in liquid diet from the 8th day gestation through 3 weeks nursing period. Offsprings, ages 1, 2 and 3 weeks, and their dams were studied. Brain weights and their total proteins were not affected by this ethanol treatment. Total enolase activity/mg protein and its isoenzymes, i.e. non-neuron-specific, hybrid and neuron-specific, (mumol/min/mg protein) in 100,000 g supernate, were significantly lower in the treated. Additionally, enolase isoenzyme transformation was delayed.


Differential effects of right-sided and left-sided orchidectomy on lateral asymmetry of LHRH cells in the mouse brain.

Luteinizing hormone-releasing hormone (LHRH) immunoreactive cells were found to be more numerous in the right side of the brain than in the left in male mice. Bilateral orchidectomy or removal of the right testis decreased the number of LHRH cells, while removal of the left testis failed to cause any changes in the number of LHRH cells. The results indicated a clear left-right difference in the neuroendocrine regulation of gonadal function in male mice.


Effects of Ca2+ antagonists and aminoglycoside antibiotics on Ca2+ current in isolated outer hair cells of guinea pig cochlea.

The effects of various Ca2+ antagonists and aminoglycoside antibiotics on the Ca2+ channel in isolated outer hair cells of the guinea pig were investigated using a whole-cell patch-clamp technique. The inhibitory action was in the order of La3+ much greater than Cd2+ much greater than Ni2+ greater than Co2+ for inorganic Ca2+ antagonists, and flunarizine = nicardipine greater than omega-conotoxin greater than methoxyverapamil = diltiazem much greater than amiloride for organic ones. Aminoglycoside antibiotics also had antagonistic effects on the Ca2+ channel.


Enhancement of acetylcholine release during REM sleep in the caudomedial medulla as measured by in vivo microdialysis.

Previous studies in our laboratory have found that muscle atonia could be triggered by two distinct areas of the medial medulla, a caudal region, corresponding to the nucleus paramedianus (NPM) and a rostral region, corresponding to the nucleus magnocellularis (NMC). The former region is responsive to acetylcholine (ACh) and the latter region is responsive to glutamate. In this study we have measured the endogenous ACh release across the sleep-wake cycle in these two areas with the microdialysis technique in unanesthetized, freely moving cats. We found that ACh release in NPM was state-dependent and was about 30% higher (P less than 0.001) during rapid eye movement (REM) sleep than during slow-wave sleep and wakefulness. However, ACh release in NMC was not selectively elevated in REM sleep. The enhancement of ACh release in NPM during REM sleep supports our hypothesis that ACh release onto cholinoceptive neurons in this area mediates the muscle atonia of REM sleep.


The neuroprotective actions of 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline (NBQX) in a rat focal ischaemia model.

The neuroprotective effects of NBQX, a selective antagonist for the AMPA/kainate subtype of excitatory amino acid receptors, were investigated in a rat focal ischaemia model, involving permanent occlusion of the left middle cerebral artery (MCA). NBQX (3, 10 or 30 mg/kg) was administered i.v. immediately after MCA occlusion and again 1 h later. The highest dose of NBQX (2 x 30 mg/kg) gave significant protection against hemispheric (24%) and cortical (27%) ischaemic damage. The lower doses of NBQX (2 x 3 or 2 x 10 mg/kg) were ineffective. No protection was seen against caudate damage for any of the doses of NBQX tested. NBQX has a t1/2 of 30 min, therefore, a second experiment was done in which a dose of 30 mg/kg was given as an i.v. bolus followed immediately by an infusion of 10 mg/kg/h for 4 h, dosing was started immediately after MCA occlusion. This dosing regimen resulted in a mean plasma level over the 4 h of 17 micrograms/ml, and significant protection against the volume of hemispheric (29%) and cortical (35%) ischaemic damage, which was slightly better than that achieved with two bolus doses of 30 mg/kg. Once again no protection was seen against caudate damage. We conclude that NBQX, an AMPA/kainate antagonist was neuroprotective in a focal ischaemia model in the rat.


Cholinergically induced REM sleep triggers Fos-like immunoreactivity in dorsolateral pontine regions associated with REM sleep.

We sought to determine the presence of Fos-like immunoreactive (Fos-LI) cells in the pontine brainstem following cholinergically induced sustained rapid-eye movement (REMc) sleep in cats. Microinjections (0.25 microliter) of vehicle (N = 2) or carbachol (2.0 micrograms/0.25 microliter; N = 4) were made into the medial pontine reticular formation. Carbachol produced a state with all the signs of natural REM sleep and with durations of 15.2-57.8 min. Compared with vehicle control animals, carbachol treated animals showed a significantly higher number of Fos-LI cells in pontine regions implicated in REM sleep generation, with longer REMc bouts associated with more Fos-LI cells than the short-duration bout. Regions with REMc-associated Fos-LI increases included: the lateral dorsal tegmental (LDT) and pedunculopontine tegmental (PPT) nuclei, where some Fos-LI cells were immunohistochemically identified as cholinergic; the locus coeruleus, where some of the Fos-LI cells were identified to be catecholaminergic; the dorsal raphe and the pontine reticular formation. These findings suggest immediate early gene activation is associated with the ubiquitous biological state of REM sleep.


Neuronal multipotentiality: evidence for network representation of physiological function.

Extracellular action potentials of single neurons in motor cortex and rectified and integrated electromyographic activity (EMG) of gastrocnemius and anterior tibialis were recorded while a monkey performed isometric ankle plantar and dorsal flexion tasks. This study determined the consistency of neuronal behaviors across different tasks. Methods characterized neuronal behaviors by determining which behavioral event within a single task, such as the appearance of the 'go' signal, force onset, or agonist and antagonist EMG onset, was best related to changes in neuronal activity. Another method compared the temporal profiles of discharge modulation across different tasks. Of 220 neurons recorded, 44 were selected because they were consistently active in the tasks. Of these, 37 were in the precentral cortex and the remaining seven were in the postcentral cortex. Only 14 of the 33 in motor cortex were consistent in their behavioral correlations. Several had multiple changes in activity within a single task that were related to different behavioral events. Half were consistent for direction of force and a third were consistent for magnitude of force. Furthermore, there was little consistency in the temporal profiles of discharge activity for all 44 neurons across tasks. Similar modulations of discharge activity among neurons in one task were different in another task. Such inconsistencies are evidence against the cardinal cell hypothesis of physiological representation. We offer a new hypothesis analogous to connectionism in parallel distributed processing.


Repeated injections of cocaine inhibit the serotonergic regulation of prolactin and renin secretion in rats.

Alterations in serotonergic function following repeated cocaine injections were examined using neuroendocrine responses to a serotonin (5-HT) releaser and 5-HT agonists. Forty-two hours following administration of cocaine (1-15 mg/kg i.p.) twice daily for 7 or 30 days, male Sprague-Dawley rats were injected with the 5-HT releaser p-chloroamphetamine (PCA; 8 mg/kg i.p.) and blood samples were collected 1 h later for radioimmunoassays of plasma prolactin, plasma renin activity (PRA) and plasma renin concentration (PRC). PCA significantly increased secretion of prolactin and renin. These responses were attenuated in rats pretreated with cocaine for 30 days. In rats receiving cocaine for 7 days, the attenuation of PCA-induced secretion of prolactin and renin was less consistently observed. To determine whether these alterations were due to pre- or postsynaptic effects, rats were injected with cocaine (15 mg/kg i.p.) twice daily for 7 days, and the neuroendocrine responses to the direct 5-HT agonists RU 24969 and m-CPP were examined, 42 h after the last cocaine injection. Pretreatment with cocaine potentiated RU 24969-induced stimulation of plasma prolactin concentration. However, cocaine did not alter the ability of m-CPP to increase plasma prolactin concentrations. The stimulation of renin secretion in response to both 5-HT agonists was not altered by cocaine pretreatment. The data suggest that repeated cocaine impairs the function of serotonergic nerve terminals that regulate these endocrine responses. Furthermore, the 5-HT receptors that mediate prolactin secretion may exhibit supersensitivity.


Vasopressin-containing neurons of the suprachiasmatic nuclei inhibit corticosterone release.

The suprachiasmatic nucleus (SCN) is the major pacemaker in the central nervous system responsible for generating circadian rhythmicity in mammals. Tracer studies show limited projections of the SCN, mainly to the paraventricular nucleus of the thalamus and paraventricular and dorsomedial nuclei of the hypothalamus, suggesting that the latter two areas may be the target areas of the SCN for controlling corticosterone release. The present results show that when infused in the paraventricular/dorsomedial nucleus of the hypothalamus femtomolar concentrations of vasopressin (VP), but not vasoactive intestinal peptide (VIP), are able to suppress elevated levels of corticosterone in SCN-lesioned animals to basal daytime values. On the other hand, infusion of the VP antagonist in the same hypothalamic area induced a sevenfold increase of basal corticosterone levels in intact animals. The SCN origin of this VP input was established in SCN-lesioned animals where no difference between the effect of infusing the antagonist or Ringer could be detected. These results imply that the SCN can influence the daily corticosterone rhythm through its VP-containing projection to the paraventricular/dorsomedial nucleus of the hypothalamus.


Maintenance and synthesis of proteins for an anucleate axon.

The anucleate (distal) segment of a crayfish medial giant axon (MGA) remains intact for months in vivo after severing the axon from its cell body, a phenomenon referred to as long-term survival (LTS). We collected axoplasm from chronic anucleate MGAs by perfusing 2-cm lengths of axons with an intracellular saline. This axoperfusate was analyzed by SDS-PAGE and silver stained. Axoperfusate proteins from intact MGAs and from chronic anucleate MGAs exhibiting LTS for up to 6 months were the same. Furthermore, immunoreactive levels of actin and beta-tubulin were similar in axoperfusates from intact and chronic anucleate MGAs. This maintenance of proteins in chronic anucleate MGAs must be due to a lack of protein degradation and/or to local protein synthesis by a source other than the cell body. To investigate local protein synthesis in vitro, we added [35S]-methionine to the extracellular saline surrounding intact and chronic anucleate MGAs. After 4- to 6-h incubations, radiolabelled proteins were detected in axoperfusates analyzed by SDS-PAGE and fluorography. The similarity between radiolabelled proteins in axoperfusates and MGA glial sheaths indicated a glial origin for the radiolabelled axoperfusate proteins. Various observations and control experiments suggested that glial-axonal protein transfer occurred by a physiological process. Glial-axonal protein transfer may contribute to the maintenance of proteins during LTS of chronic anucleate MGAs.


Ventral pontine catecholaminergic pathway mediates the vasopressin response to splanchnic osmostimulation in conscious rats.

This study examines the role of catecholamines, cell bodies and fibers of passage within the subcoeruleus area (subLC) in the arginine vasopressin (AVP) response to splanchnic osmotic stimulation and hemorrhage. Bilateral chemical lesions were induced into the subLC, approximately 1 mm ventral to the locus coeruleus (LC), using 6-hydroxydopamine (6-OHDA) and ibotenic acid to selectively destroy catecholaminergic components and cell bodies, respectively. Vehicle and 5,7-dihydroxytryptamine (5,7-DHT) injections into the subLC area, 6-OHDA injections into the LC, as well as systemic desipramine pretreatment, were performed as controls for the possible non-specific effects of the lesions. Seven and 8 days later, plasma AVP level, plasma osmolality, mean arterial pressure and heart rate were measured following either gastric infusion of hypertonic (598 mOsm/kg; 2 ml/4 min) or isotonic (290 mOsm/kg) saline or a mild hemorrhage (2.5 ml/300 g) in conscious rats with indwelling tail artery catheters and naso-gastric tubes. 6-OHDA injections into subLC reduced the AVP response to the osmotic stimulation by 62.3% (P less than 0.01), as compared to vehicle-injected controls. These same rats demonstrated a normal AVP response to hemorrhage implying a specificity of the disrupted pathway. All controls confirmed that the effects of the 6-OHDA were due to specific action on noradrenergic components within the subLC area. Ibotenic acid lesions in the subLC did not significantly decrease the AVP response, demonstrating that mainly fibers and not cell bodies in this region are part of the pathway. 6-OHDA injections just anterior to the LC, where the dorsal noradrenergic bundle (DNAB) forms, reduced the AVP secretion due to hemorrhage by 77.0% (P less than 0.05), but had minor effects on the response to osmotic stimulation. These results indicate that catecholaminergic fibers travelling primarily within the subLC, in the ventral noradrenergic bundle (VNAB), carry splanchnic osmotic input to the hypothalamus, whereas the DNAB may mediate the AVP response to hemorrhage.


Disappearance of circadian rhythms in Parkinson's disease model induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine in dogs.

Administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) to dogs produces clinical, pathological and neurological features in dog resembling human Parkinson's disease. Using this animal model, we studied the changes in diurnal rhythms of urine volume, creatinine in urine, and vasopressin, aldosterone and renin activity in plasma. Before MPTP treatment, urine volume showed a peak between 17.00 and 1.00 and plasma vasopressin concentration also showed a clear circadian rhythm with a peak at 13.00 and a minimum level at 5.00. Two weeks after MPTP treatment (2.5 mg/kg i.v.), the rhythm of urine volume disappeared and that of vasopressin became less clear. Plasma renin activity increased 2 and 4 weeks after MPTP treatment. The increase was, however, not enough to change the concentration of plasma aldosterone. We examined the effect of L-3,4-dihydroxyphenylalanine (levodopa), on the circadian pattern of urine volume and vasopressin attenuated by MPTP. Levodopa (4 mg/kg/day) was administered orally every day from the first week after MPTP treatment. The circadian rhythms of urine volume and vasopressin reappeared within one week after the start of levodopa administration.


Intraretinal xenografts of differentiated human retinoblastoma cells integrate with the host retina.

We report on the successful use of chemically modified Y79 human retinoblastoma cells for intraretinal xenografting into damaged adult mammalian eyes. Y79 cells were exposed in vitro to retinoic acid/butyrate to induce differentiation. Using a multisite transplantation method, the suspension was injected into the subretinal space of Fischer 344 rats. The survival, integration, and differentiation potential of these cells was studied, following their return to the intraocular milieu from which the progenitor cells originated. The grafted cells survived and differentiated into immature photoreceptor elements in the subretinal and intraretinal locations, as multiple clusters of rosette-forming cells intimately attached to the host neuroretina. The differentiation process included development of synaptic connectivity of the ribbon type with the surrounding neuropil. No signs of renewed cell division were found within grafts performed on 42 rat eyes, and there was no indication of cell-mediated host reaction against the transplants. This study indicates that tumorigenicity can be suppressed in mitotically arrested Y79 cells, and that these cells are capable of undergoing differentiation in vivo. This provides evidence of the remarkable differentiation properties of human retinoblastomas while indicating that Y79 cells may ultimately be able to substitute for fetal cells in experimental retinal transplantation.


The effects of lesions of the habenular nuclei on the development of sensitization to the behavioral activational effects of repeatedly administered morphine in the rat.

The effects of lesions of the habenular nuclei on the development of sensitization to the behavioral activational effects of morphine (MOR), administered repeatedly either systemically or directly into the ventral tegmental area (VTA) were examined. Lesions of the habenular nuclei blocked the early-appearing sedative effects and enhanced the later-appearing locomotor activational effects seen after systemic injections of MOR (10 mg/kg, i.p.). Habenular lesions did not potentiate the development of sensitization to the locomotor-activational effects seen with the repeated, systemic administration of MOR. The bilateral injection of MOR (5.0 micrograms/0.5 microliter/side) directly into the VTA of animals with habenular lesions resulted in the performance of stereotyped behaviors that appeared as early as the second MOR exposure and remained at high levels with repeated MOR treatment. The stereotyped behavior shown by lesioned animals did not appear to interfere with the acute locomotor activational effects of intra-VTA MOR nor the development of sensitization to these effects when it was administered repeatedly. These results are in agreement with previous research suggesting that by disinhibiting the dopamine (DA) systems, habenular lesions enhance the acute behavioral activational effects of MOR. The results also suggest that the habenular nuclei do not control the changes in the response of the DA systems underlying the development of sensitization to the locomotor-activating effects of MOR when administered repeatedly.


The response of the 5-hydroxyindole oxidation current to noxious stimuli in the spinal cord of anesthetized rats: modification by morphine.

The effects of cutaneous noxious heating and of systemic morphine on serotonergic activity in the spinal cord were examined in anesthetized rats. An oxidation current of 5-hydroxyindole signal was seen at 280-300 mV with differential normal pulse voltammetry. Noxious heat stimuli produced a mean signal increase over control values of 15.5 +/- 3.4% at 52 degrees C, and 7.2 +/- 5.5% at 45 degrees C. These increases lasted for 5-10 min. Non-noxious stimuli (37 degrees C) did not affect the 5-hydroxyindole signal. Morphine (0.5, 2.0 and 5.0 mg/kg, i.p.) in the absence of cutaneous stimulation did not change the signal significantly. Systemic morphine alone did not significantly modify the 5-hydroxytryptamine (5-HT) metabolism, as observed in in vivo voltammetry, in the spinal cord of anesthetized rat. However, a low dose of morphine (0.5 mg/kg, i.p.) attenuated the increase in the signal modified by noxious stimuli, and high doses (2.0 or 5.0 mg/kg, i.p.) enhanced it. Both effects of morphine were antagonized by naloxone (0.5 mg/kg, i.v.). It is likely that morphine with noxious stimuli modify the sensitivity of serotonergic descending inhibitory system. It is concluded that noxious heating of the skin increases the 5-HT metabolism in the spinal cord of anesthetized rats and that systemic administration of morphine modulates this 5-HT metabolism.


Abnormal profiles of polyunsaturated fatty acids in the brain, liver, kidney and retina of patients with peroxisomal disorders.

The polyunsaturated fatty acid (PUFA) composition of the brain was studied in 8 patients with Zellweger's syndrome (ZS), 3 with neonatal adrenoleukodystrophy (NALD), one with bifunctional enzyme deficiency (BED), one with X-linked adrenoleukodystrophy (X-ALD), and one with adrenomyeloneuropathy (AMN). The PUFA composition of the liver, kidney and retina was studied in 8, 6 and 1 patients with ZS, respectively. An infant with NALD and a child with rhizomelic chondrodysplasia punctata (RCDP) were also studied for the PUFA composition of the liver. The liver and kidney of the patient with X-ALD and the liver of the patient with AMN were included in the study. The fatty acid values in the peroxisomal patients were compared with control data obtained in the normal developing brain (38 cases), liver (9 cases), kidney (7 cases) and retina (16 cases). The brain of a patient with metachromatic leukodystrophy (MLD) and the liver of a child with Krabbe's disease (KD) were also studied for comparison. The most constant and severe abnormality in all the peroxisomal patients was a drastic decrease in the total amount of docosahexaenoic acid (22:6 omega 3), especially in the brain. The other product of delta 4-desaturation, 22:5 omega 6, was generally decreased in the brain, liver and kidney of the ZS patients, but very much increased in the brain of two patients with NALD. The 22:6 omega 3/22:4 omega 6 ratio, which remains quite constant throughout normal brain development, was consistently decreased in the peroxisomal brain, in ZS as well as in NALD. This study confirms that, in classical Zellweger's syndrome, the two products of delta 4-desaturation are affected. In contrast, in neonatal adrenoleukodystrophy the deficiency is probably restricted to the omega 3 product of delta 4-desaturation, docosahexaenoic acid, especially in the brain, while the other product, 22:5 omega 6, is either normal or increased, perhaps in an attempt to compensate for the 22:6 omega 3 deficiency in brain membranes.


Evidence that long-term estrogen treatment disrupts opioid involvement in the induction of pituitary LH surge.

Recent evidence suggests that a decrease in the inhibitory opioid influence is a necessary hypothalamic neural event in the preovulatory and ovarian steroid-induced luteinizing hormone (LH) surge. Whether shifts in ovarian steroidal milieu disrupts this neural event is not known. Therefore, the effects of short-term (3 days) and long-term (13 or 17 days) estradiol 17 beta (E2) exposure on spontaneous, naloxone (NAL) and progesterone (P)-induced LH surges were assessed in ovariectomized (ovx) rats. Two weeks after ovariectomy, rats received subcutaneous Silastic capsules filled with crystalline E2. In rats exposed to E2 for 3 days and infused with saline intravenously between 11.00-14.00 h, plasma LH rose significantly at 17.00 h. NAL infusion between 11.00-14.00 h in these rats to decrease the inhibitory opioid influence, advanced both the onset of LH rise and amplified the secretion of LH in the afternoon. Continuation of E2 exposure for 13 days produced no deleterious effects on either the spontaneous or NAL-induced augmentation in LH responses. However, uninterrupted E2 exposure for 17 days abolished the spontaneous afternoon LH rise and drastically diminished the ability of NAL to advance and amplify the LH response. In the next experiment, we evaluated the effect of P injection (2 mg/rat) at 11.00 h on the afternoon LH release in rats similarly exposed to E2 for either 3 or 17 days. In rats exposed to E2 for 3 days, the P-induced LH release was significantly greater than that observed after saline or NAL infusion.(ABSTRACT TRUNCATED AT 250 WORDS)


Role of glutathione in repair of free radical damage in hippocampus in vitro.

Depletion of glutathione (GSH), an intrinsic antioxidant, increases vulnerability to free radical damage in a number of cell systems. This study investigates the role of GSH in limiting electrophysiological damage and/or recovery from free radical exposure in slices of guinea pig hippocampus. Synaptic potentials (PSPs) and population spikes (PSs) were recorded from field CA1. Free radicals were generated from 0.006% peroxide through the Fenton reaction. Analysis of the input-output curves showed that peroxide treatment decreased PSPs and impaired ability of the PSPs to generate PSs as previously reported. Recovery was nearly total within a half hour. Treatment with 5 mM buthionine sulfoximine (BSO) for 2 h depleted hippocampal GSH to 79.2% of control values. The extent of free radical damage was not increased. Recovery, however, was only partial. GSH was further depleted by oxidation with diamide or covalent bonding with dimethyl fumarate (DMF) immediately before and during the peroxide treatment. Neither diamide nor DMF treatment in BSO-incubated tissue enhanced peroxide-induced electrophysiological deficits. Following these treatments, however, tissue showed little recovery from free radical damage. We conclude that glutathione is essential for repair processes in hippocampal neurons exposed to oxidative damage.


Fos expression in intrastriatal striatal grafts: regulation by host dopaminergic afferents.

Previous studies have shown that transplants of fetal striatum, implanted into the ibotenic acid-lesioned striatum of adult rats, become innervated from the host nigrostriatal dopamine (DA) pathway. In the present study we have used DA-receptor-mediated expression of the Fos protein (i.e. the product of the immediate-early c-fos gene) as a cellular marker for functional dopaminergic host-graft interactions in the striatal grafts. Amphetamine (5 mg/kg; 2 h) induced Fos-like immunoreactivity in clusters of cells located mainly within the DARPP-32-positive areas within the transplants, i.e. within the striatum-like graft compartment which is preferentially innervated by the host DA afferents. As in the normal striatum, this effect was largely, although not completely, abolished by a 6-hydroxydopamine lesion of the ipsilateral nigrostriatal DA pathway. Apomorphine (0.25 mg/kg; 2 h) had no detectable effect in grafts with an intact host DA system. Two to 3 weeks after a 6-OHDA lesion of the host DA pathway (i.e. a time sufficient for DA receptor supersensitivity to develop), apomorphine-induced extensive Fos-activation selectively within the DARPP-32-positive areas of the graft. The magnitude of the response was similar to that seen in the DA-denervated host striatum. Dual Fos/DARPP-32 immunostaining revealed that the activated graft neurons were, at least in part, DARPP-32-positive. In intrastriatal grafts of fetal neocortical tissue, which were studied for comparison, the amphetamine- and apomorphine-induced effects on Fos expression were much smaller and similar to that seen in the DARPP-32-negative, non-striatal compartment within the striatal grafts.(ABSTRACT TRUNCATED AT 250 WORDS)


Behavior of axons, Schwann cells and perineurial cells in nerve regeneration within transplanted nerve grafts: effects of anti-laminin and anti-fibronectin antisera.

Wistar rats (close cloned strain) were used to investigate the effect of endogenous laminin and fibronectin on axons, Schwann cells and perineurial cells in the regenerating peripheral nervous system (PNS). Sciatic nerve grafts obtained from donor rats were frozen, thawed and treated with rabbit anti-rat laminin or anti-fibronectin antiserum. Control grafts were treated with normal rabbit serum alone. One cm long portions of the sciatic nerve of the recipient rats were replaced with grafts. At 15 days after transplantation the number of regenerated axons in the laminin- and fibronectin-depleted grafts was half of that in the control. The growing axons in the laminin-depleted grafts did not recognize the basal lamina scaffolds (BLS) remaining in the basal lamina tubes, while in the control and fibronectin-depleted grafts 90% or more of axons grew inside the BLS. Elongation of axons always preceded migration of Schwann cells with the latter subsequently adhering to and wrapping around the former. Perineurium-forming fibroblastic cells recognized the combination of axons and Schwann cells and formed perineurial fasciculi around them. These fibroblastic cells did not recognize empty BLS but responded to them only when fibronectin was depleted. Macrophages sometimes closely faced the naked axons which elongated outside the BLS. These results suggest that in the early stages of nerve regeneration endogenous laminin and fibronectin not only regulate the growth of regenerating nerve fibers, but also exert a positive influence on perineurial cells and macrophages, both of which play important roles in nerve tissue injury and repair.


Testosterone reverses a senescent decline in extrahypothalamic vasopressin mRNA.

The biosynthetic activity of extra-hypothalamic vasopressin (VP) neurons in the bed nucleus of the stria terminalis (BNST) is regulated by gonadal steroids. These neurons have also been implicated in a number of behaviors that are impaired in aging. We previously reported that VP mRNA labelling in the BNST is decreased in senescent rats. We hypothesized that the age-related decrease in VP mRNA labelling is due to the decline in circulating testosterone (T) levels in aged animals. T or saline was administered peripherally for 1 month in physiologic or superphysiologic doses to 3 month old or 24 month old Fischer 344 male rats. In situ hybridization and quantitative autoradiography for VP mRNA in the BNST were performed using a 48-base 35S-labelled oligonucleotide probe. Administration of T completely reversed the decline in VP mRNA labelling in the aged animals. Superphysiologic T further increased VP gene expression in both age groups. These data are consistent with a previous report of T-induced increase in VP immunoreactive fiber density in other extrahypothalamic regions of the brain in aged rats. This study offers further evidence that alterations in the hormonal milieu may play an important role in modulating neuronal biosynthetic activity in senescence.


Auto- and cross-correlation analysis of subthalamic nucleus neuronal activity in neostriatal- and globus pallidal-lesioned rats.

Statistical analyses (autocorrelation and first-order interstimulus interval) were conducted on the spontaneous activity of over 420 subthalamic neurons recorded in 5 groups (control, large globus pallidus kainic acid lesion, partial globus pallidus kainic acid lesion, partial globus pallidus ibotenic acid lesion and neostriatal lesion) of anesthetized rats. Cross-correlation and peristimulus time histogram (to frontal motor cortex stimulation at 0.7 mA) analyses were conducted on pairs (n = 58) of subthalamic neurons recorded simultaneously on a single microelectrode. Lesion of the globus pallidus increased spontaneous firing rate as compared to controls and shifted the pattern of spontaneous activity from either a regular or irregular pattern to a markedly bursting pattern. Neostriatal lesion reduced firing rate and reduced the likelihood of highly regular firing. In control, neostriatal and partial lesioned animals, approximately 1 in 3 pairs of neurons showed correlated firing. The correlations were joint increased probabilities of firing over intervals of 200-400 ms, suggesting a shared excitatory input. No short-interval (less than 10 ms) correlations were seen. Large globus pallidus lesion increased the likelihood of correlated firing (12 of 16 pairs). In all groups of animals the peristimulus time histograms (PSTHs) to motor cortex stimulation were more similar than would be expected by chance and pairs of neurons showed the same increases in response following globus pallidus lesion. Thus adjacent neurons share common cortical inputs and responsiveness to those inputs. These changes indicate that the globus pallidus influences the spontaneous firing rate and pattern of subthalamic neurons as well as the degree of correlated firing of adjacent neurons.


Biochemical characterization of independent olfactory receptor sites for 5'-AMP and taurine in the spiny lobster.

To understand the initial events in chemosensory transduction (i.e. binding of odorants to olfactory receptor sites), we have utilized a radioligand-receptor binding assay on a tissue preparation that is enriched in dendritic membrane from olfactory receptor cells in the spiny lobster Panulirus argus. Radioligands used were tritiated adenosine 5'-monophosphate (AMP) and taurine, which are two of the most excitatory compounds for spiny lobsters. Our results indicate the existence of two independent types of binding sites--a taurine binding site and an AMP binding site. For both the taurine and AMP binding sites, odorant binding is rapid, reversible, and saturable. KD values for the taurine and AMP binding sites are 2.3 and 2.0 microM, respectively, and Bmax values are 159 and 3.2 fmol/micrograms protein, respectively. The fact that the specificity, affinity, and independence of these two binding sites as defined in these biochemical studies are in agreement with those from electrophysiological studies suggests that these binding sites are olfactory receptor sites involved in sensory transduction.