Distribution of SNAP25, VAMP1 and VAMP2 in mature and developing deep cerebellar nuclei after estrogen administration.

Synaptosomal-associated protein of 25kDa (SNAP25), vesicle-associated membrane protein 1 (VAMP1) and 2 (VAMP2) are components of soluble N-ethylmaleimide-sensitive fusion attachment protein receptors (SNARE) complex which is involved in synaptic vesicle exocytosis, a fundamental step in neurotransmitter release. SNARE expression in cerebellum correlates with specific neurotransmitter pathways underlying synaptic diversification and defined synaptic properties. In this study we firstly characterized the distribution of SNAP25, VAMP1 and VAMP2 in the nerve terminals of a defined cerebellar region, the deep cerebellar nuclei (DCN), of adult and newborn rats. Then, given the pivotal role of estradiol (E2) in the synaptic organization of the cerebellar circuitry in early postnatal life, we examined whether administration of E2 in the newborn DCN affected synaptic density and changed the distribution of the presynaptic proteins SNAP25, VAMP1 and VAMP2, together with post synaptic density protein 95 (PSD95). Results showed that: (1) distribution of SNAP25, VAMP1 and VAMP2 in adult DCN differs significantly from that found in newborn DCN; (2) administration of E2 in the newborn DCN affected synaptic density and also changed the distribution of the pre- and postsynaptic proteins. The differential distribution of SNAP25, VAMP1 and VAMP2 in nerve terminals of adult and newborn rats may correlate with specific stages of neuronal phenotypic differentiation. The effects of E2 on SNAP25, VAMP1, VAMP2, PDS95 and synaptic density suggest that pre- and postsynaptic proteins are under estrogenic control during development and that synaptic maturation can also be related with the activity of this steroid.

Immunohistochemical localisation and molecular expression of the steroidogenic enzyme cytochrome P450 17α-hydroxylase /C(17,20)-lyase in the vestibular nuclei of adult male rats.

Many biologically active neurosteroids, including dehydroepiandrosterone (DHEA), are synthesised in the brain. DHEA is a potent endogenous modulator of several neuronal functions, and alterations of DHEA are correlated with various neurobiological deficits. The cytochrome P450 17α-hydroxylase/C(17,20)-lyase (P450C(17) ) plays a pivotal role in the synthesis of DHEA from pregnenolone and progesterone. We investigated the immunohistochemical localisation and molecular expression of P450C(17) in the superior, lateral, medial and inferior vestibular nuclei (VCN) of adult male rats by western blotting and indirect immunofluorescence analysis. Immunoreactive P450C(17) was widely distributed in all VCN and the expression of P450C(17) was confirmed by western blot analysis. The present study demonstrates, for the first time, the presence and anatomical distribution of P450C(17) in the VCN. Given that neurosteroids can modulate neuronal activities in the medial vestibular nucleus, DHEA synthesised in the VCN may play an important role in the control of specific activities at this level.

Hypoglossal nucleus projections to the rat masseter muscle.

We investigated in the rat whether hypoglossal innervation extended to facial muscles other than the extrinsic musculature of the mystacial pad. Results showed that hypoglossal neurons also innervate the masseter muscle. Dil injected into the XII nucleus showed hypoglossal axons in the ipsilateral main trunk of the trigeminal nerve. After Gasser's ganglion crossing, the axons entered into the infraorbital division of the trigeminal nerve and targeted the extrinsic muscles of the mystacial pad. They also spread into the masseter branch of the trigeminal nerve to target the polar portions of the masseter muscle spindles. Retrograde double labelling, performed by injecting Dil into the pad and True Blue into the ipsilateral masseter muscle, showed labelled hypoglossal neurons in the medio-dorsal portion of the XII nucleus. The majority of these neurons were small (15-20 microm diameter), showed fluorescence for Dil and projected to the mystacial pad. Other medium-size neurons (25 microm diameter) were instead labelled with True Blue and projected to the masseter muscle. Finally, in the same area, other small hypoglossal neurons showed double labelling and projected to both structures. Functional hypotheses on the role of these hypoglossal projections have been discussed.

Thyroid hormone action: nongenomic modulation of neuronal excitability in the hippocampus.

Years of effort have failed to establish a generally-accepted mechanism of thyroid hormone (TH) action in the mature brain. Recently, both morphological and pharmacological evidence have supported a direct neuroactive role for the hormone and its triiodinated metabolites. However, no direct physiological validation has been available. We now describe electrophysiological studies in vivo in which we observed that local thyroxine (T4) administration promptly inhibited field excitatory postsynaptic potentials recorded in the dentate gyrus (DG) with stimulation of the medial perforant pathway, a result that was found to be especially pronounced in hypothyroid rats. In separate in vitro experiments, we observed more subtle but statistically significant responses of hippocampal slices to treatment with the hormone. The results demonstrate that baseline firing rates of CA1 pyramidal cells were modestly reduced by pulse-perfusion with T4. By contrast, administration of triiodothyronine (T3) was often noted to have modest enhancing effects on CA1 cell firing rates in hippocampal slices from euthyroid animals. Moreover, and more reliably, robust firing rate increases induced by norepinephrine were amplified when preceded by treatment with T3, whereas they were diminished by pretreatment with T4. These studies provide the first direct evidence for functional, nongenomic actions of TH leading to rapid changes in neuronal excitability in adult rat DG studied in vivo and highlight the opposing effects of T4 and T3 on norepinephrine-induced responses of CA1 cells studied in vitro.

Hypoglossal nuclei participation in rat mystacial pad control.

Recently, we showed that extra-trigeminal axons, originating from the hypoglossal nucleus, travel with the infraorbital division of the trigeminal nerve (ION), which is known to innervate the rat mystacial pad. Dil was monolaterally injected into the rat XII nucleus to analyse the peripheral distribution of hypoglossal axons to the mystacial pad, to evaluate their involvement in facial sensory-motor control. Electromyographic responses of mystacial pad motor units to electrical stimulation of the ION were recorded, along with the evoked responses to electrical stimulation of the ipsilateral XII nucleus. The results showed that hypoglossal axon terminals target the ipsilateral extrinsic musculature of the mystacial pad, but they do not have any contact with the intrinsic muscles. ION electrical stimulation increased electromyographic activity in the ipsilateral pad extrinsic muscles, even following VII nerve transection. Hypoglossal nucleus electrical stimulation induced field potentials and monosynaptic responses in the same motor units that persisted even following VII nerve transection, these disappearing after cooling the ION. We suggest that the small hypoglossal neurons projecting to the extrinsic musculature of the mystacial pad are part of a hypoglossal-trigeminal loop that participates in the sensory-motor control of the rat vibrissae system.

Quantification of Spinning bike performance during a standard 50-minute class.

Spinning is a type of indoor fitness activity performed on stationary bikes by participants who pedal together to the rhythm of music and the motivating words of an instructor. Despite worldwide popularity of this type of recreational activity, to date there have been few, mainly non-scientific, studies of the impact of spinning on metabolic, respiratory, and cardiovascular functions. The main aim of this study was to evaluate a number of metabolic and cardiovascular variables during a standard 50-min class performed by Spinning instructors of both sexes: six males (age 30 +/- 4.8 years, body mass index 24 +/- 2.5 kg x m(-2); mean +/- s) and six females (age 34 +/- 6.3 years, body mass index 21 +/- 1.9 kg x m(-2)). The mean power output, heart rate, and oxygen uptake during the performance were 120 +/- 4 W, 136 +/- 13 beats x min(-1), and 32.8 +/- 5.4 ml x kg(-1) x min(-1) respectively for males, and 73 +/- 43 W, 143 +/- 25 beats x min(-1), and 30 +/- 9.9 ml x kg(-1) x min(-1) respectively for females. Analysis of individual performances showed that they were compatible with physical exercise that ranged from moderate-to-heavy to very heavy, the latter conditions prevailing. The results show that this type of fitness activity has a high impact on cardiovascular function and suggest that it is not suitable for unfit or sedentary individuals, especially the middle aged or elderly, who are willing to begin a recreational physical activity programme.

Effect of aluminum consumption on the vestibulo-ocular reflex.

The effects of chronic exposure (90 days) to Aluminum chloride (AlCl(3)) were analyzed in 3, 10 and 24 month old male rats (n=270) by investigating the function of the VOR (vestibulo-ocular reflex) in correlation with Aluminum (Al) concentrations in blood and brain. The VOR was chosen and tested in basal conditions (pre-exposure measures) and during the continuous administration of three different concentrations of AlCl(3) (0.5, 1, 2 g/l in drinking solution): the control animals being exposed to NaCl (0.125, 0.25 and 5 g/l in drinking solution). Results showed that LOAEL (Lowest Observed Adverse Effect Level) was 43.1+/-11.4 mg Al/kg-day. At this concentration the Al induced significant VOR impairment in all exposed rats, regardless of animal age. Neuroanatomical analysis showed that VOR impairment was not dependent on astrocyte damage nor evidences of amyloid deposits in the exposed rats was found. Significant changes of Al concentrations were observed in nervous tissue, while its concentration in whole blood was unaffected. Furthermore, results show that it is possible to identify an individual neurotoxic threshold for each animal and therefore hypothesize the clinical use of the VOR test for the evaluation of individual risk toxicity to chronic Al exposure.

Sudden death in epilepsy: an experimental animal model.

INTRODUCTION: The physiopathogenetic mechanisms possibly involved in sudden unexplained epileptic death (SUDEP), were investigated in the hemispherectomized rat. METHODS: For this purpose, paroxysmal activity, vagal nerve firing, systemic blood pressure (BP), pulmonary artery pressure, and ECG were simultaneously recorded in an experimental animal model of epilepsy. Recordings were performed in basal conditions and during paroxysmal activity induced by topical application of penicillin-G at hypothalamic and mesencephalic level. During the experiment were also performed hemogas analysis and at end, samples of lung tissue were processed for histology. RESULTS: Activation of hypothalamic (HEF) and mesencephalic (MEF) epileptic foci induced a significant increase of spontaneous vagal nerve firing that was strictly correlated to ECG impairments and hypotension. When paroxysmal activity extinguished, vagal nerve activity and cardiovascular parameters returned to basal conditions. However, in 25% of the animals, co-activation of HEF and MEF always triggered a vagal hypertone which was temporally correlated to cardiac arrhythmias, but also to hyperkalemia, acidosis, pulmonary hypertension and to animal death. Histological control in lungs of deceased animals showed an alveolar and perivessel oedema with an oedematous infiltration in the alveolar and bronchial spaces and mucous secretion. During ictal activity, comparison between survived and deceased animals showed significant differences in the incidence of ECG impairment of pulmonary artery pressures, pO2, and pCO2 pressures, and [K+], [HCO3-], and [pH], concentrations. DISCUSSION: A possible explanation of the above observations is discussed in relationship to SUDEP physiopathogenesis.

Frequency-dependent LTP/LTD in guinea pig Deiters' nucleus.

Synaptic plasticity was studied in the lateral vestibular nuclei (LVN) of the guinea pig in vivo. High frequency stimulation (HFS) of increasing or decreasing frequencies was applied to the ipsilateral vestibular nerve. Vestibular field potentials (VFPs) and extracellular single unit activity evoked in the LVN by electrical stimulation of the ipsilateral vestibular nerve, were analyzed before and after the application of different protocols of HFS. Results show that the monosynaptic component of the VFPs undergo long-term potentiation (LTP) with stimulation of 100 Hz applied for 20 s lower frequencies, applied for shorter periods, induce only a transient post-tetanic potentiation. This potentiation, although long lasting, is not permanent since it is susceptible of a reversal or cancellation by opposite patterns of HFS that determine a depression or depotentiation of the previously acquired potentiation. The results demonstrate that the plasticity phenomena that take place at the level of the LVN neurons are not steady but undergo continuous adjustment of their sign and gain depending on the variable flow of vestibular information that reach the nuclei from the labyrinthine receptors.

Neurotoxic effect of lead at low concentrations.

The effects of lead exposure at low concentrations were evaluated by studying the post-rotatory nystagmus (PRN) in two groups of rats exposed for 3 months to 50 parts per million (ppm) of sodium acetate and 50 ppm of lead acetate, respectively, in the drinking water. Only animals treated with lead acetate showed changes of the PRN parameters which were significantly related to the concentration of lead in the blood and in brain structures. The patterns of PRN responses were characterized and classified into four types: progressively inhibitory (40%), prematurely inhibitory (25%), late inhibitory (25%), and excitatory-inhibitory (10%). No alterations of the PRN parameters were observed in the animals treated with sodium acetate. The results show that exposure to lead, even at low concentrations, impairs both sensory and motor functions. The findings also point out that the vestibular system and brain stem structures which generate and control the PRN represent targets of the action of this heavy metal. Finally, the results indicate that the evaluation of the vestibulo-ocular-reflex can provide a test suited for the screening of the neurotoxic effects of lead even in the absence of clinical signs typical of lead intoxication.

Autonomic nervous system activity and life threatening arrhythmias in experimental epilepsy.

In the present study the possible derangement of the autonomic system and its influence in life threatening arrhythmias were analysed during paroxysmal activity. In hemispherectomized rats a paroxysmal activation of the hypothalamic and mesencephalic cardioarrhythmogenic triggers was performed by topical application of penicillin-G. Blood gas parameters and electrical activity of the thalamus, hypothalamus, vagal nerve fibre, ECG and arterial blood pressure were simultaneously monitored in basal conditions and repeated after the appearance of paroxysmal activity. Temporal correlation analysis was carried out. Results showed that during activation of these triggers, the spontaneous vagal nerve fibre activity significantly increased and triggered the appearance of cardiac arrhythmias which could become life threatening and induce animal death when blood gas and electrolytic parameters were simultaneously impaired. These experiments suggest that fatal evolution of the heart impairment is related not only to an autonomic cardiac trigger, but also to a concomitant metabolic derangement, which most likely shares the same autonomic origin.

Disposition of propofol between red blood cells, plasma, brain and cerebrospinal fluid in rabbits.

The disposition of propofol in the blood and brain of New Zealand rabbits was studied in three groups of six rabbits. One group received a single anaesthetic dose; a second group received a 1-h infusion; and a third group was studied after the rabbits were judged to have recovered from a 1-h infusion. There was a high concentration of propofol in the red blood cell fraction and in the brain, however, the red blood cell concentration largely exceeded the one found in the brain in all groups of animals. This is consistent with the high fat solubility of diisopropylphenol. The possible effects of propofol sequestered in red blood cells is discussed.

Assessment of the metals contamination of the low Sebou sediments (Morocco).

The heavy metals contamination of the lower Sebou sediments was studied to determine the average degree of contamination and to assess the extent of anthropogenic contamination. The spatial and temporal distribution of these metals and the results of the contamination indices showed very serious metallic contamination principally of Cr, Pb and Hg. However accurate analysis of the situation in the river mouth was complicated by the fact that the metals could be moved by the tides and currents.

The impact of lead pollution on the environment of Rabat-Sale' (Morocco).

There are numerous possible vectors of industrial lead poisoning in unprotected work environments and the areas surrounding industrial sites. In the Rabat-Salé region, ceramics is the industry which uses most lead. Galena (PbS) is used in the fabrication of some types of pottery, and other lead-based compounds are used as glaze. A second source of lead pollution is the production of some fuels, in which lead is used as an anti detonator. In order to evaluate the impact of lead contamination on the environment, our investigation is focused on the lead concentration in the atmosphere, soil and vegetation at five stations in the Rabat-Salé region. We found concentrations of lead higher than those published for any other country in the atmosphere and vegetation near the ceramics factories of Rabat-Salé. These concentrations decreased significantly as we moved away from urban areas.

Epileptic discharge of cortical, subcortical and spinal neurons in penicillin induced experimental epilepsy.

The sensitivity and electrophysiological patterns of paroxysmal activity induced in different brain structures by topical application of penicillin-G were evaluated in the rat. Recordings were carried out in five groups of animals, in telencephalon, diencephalon, mesencephalon, rombencephalon and spinal cords. The following analysis were carried out: frequency distribution histograms, latency and time course duration of paroxysmal activity, duration and amplitude of epileptic bursts. The results obtained showed that the nervous structures tested with penicillin-G had a different epileptogenic sensitivity and response pattern which significantly changed along the cerebral cortex-spinal cord axis. The highest epileptic sensitivity was observed in somatosensory cortex (SI) at 500-600 microns depth; in the other cortical layers, a significant lenghtening in latency was observed. Among the other structures, the spinal cord seemed to be the most sensitive target to the epileptogenic action of penicillin-G, whereas in the remaining structures, sensitivity significantly decreased in rostro-caudal direction. As far as the features of the paroxysmal activity are concerned, significant differences among tested structures were observed. In particular, within the SI cortex, the main differences were represented by the gradual increase in burst frequency and voltage from the surface to the IVth layer and by their subsequent decrease in deeper layers (V-VI). In the diencephalon, the paroxysmal activity was similar to that observed in more superficial and deeper cortical layers even though epileptic bursts showed a lower amplitude. Mesencephalon and rombencephalon displayed a paroxysmal activity with a distinctive feature, characterized by long lasting bursts of low amplitude, although bulbar outbursts showed a shorter duration than the mesencephalic ones. In the spinal cord, the epileptiform activity displayed a different paroxysmal pattern, characterized by the longest duration and the highest amplitude. The different sensitivities of the investigated brain structures to penicillin-G and the characteristics of the induced paroxysmal activity have been extensively discussed.

Beta-endorphin and cortisol levels in plasma and CSF following acute experimental spinal traumas.

beta-endorphin and cortisol were measured in cerebrospinal fluid (CSF) and plasma by radioimmunological method (RIA) in two groups of rabbits with spinal cord traumatic injuries at cervical and lumbar levels, respectively with and without concomitant spinal shock and arterial hypotension, and in a group of sham operated animals as controls. The two groups with spinal lesions displayed a significant beta-endorphin increase in CSF, whereas the cortisol level remained unchanged both in the spinal traumatized rabbits and in controls. Both the opioid and the cortisol concentration rose significantly in plasma in all three groups and in particular resulted significantly higher in the cervical traumatized group where spinal trauma was associated with spinal shock and hypotension. However, no significant difference was found when beta-endorphin concentrations in plasma were compared between the sham operated animals and the spinal lumbar traumatized animals without concomitant spinal shock. The results seem to suggest that the beta-endorphin increase in CSF is related to the nervous tissue lesion, while its increase in plasma, like that of cortisol, is due to surgery or other stress factors inherent in the experiment. This independent behaviour of beta-endorphin in plasma and in CSF suggests its different origin in these two compartments.

Olfactory influence on tongue activity.

Recent findings have shown that olfactory stimulation by brief puffs of air odorized with amyl acetate induces several patterns of response in rabbit hypoglossal neurons. It has been argued that the functional role of the olfactory input may be the modulation of tongue muscular tone during the oral phase of digestion. In the present research, the peripheral effect of olfactory-hypoglossal modulation was analyzed. Both the spike traffic along the fibers of the hypoglossal nerve and the electromyographic activity of single tongue muscles (genioglossus, styloglossus, superior longitudinal and hyoglossus) were recorded before, during and after olfactory stimulation. Results showed that brief puffs of air odorized with amyl acetate induced a significant change in the efferent volleys along the hypoglossal nerve, as well as a substantial modulation of tongue muscle activity. Olfactory stimulation induced a significant increase in the spontaneous activity of both type I and type II genioglossal fibers; excitation followed by inhibition both in tonic and phasic styloglossal fibers; excitatory responses in tonic and phasic superior longitudinal fibers and short-lasting excitatory responses in the hyoglossal fibers. The diverse patterns of activation of the tested muscle and the significant differences between fibers, tonically or phasically controlled by the XIIth neurons, indicate that olfaction may be strongly involved in tongue reflex regulation. Different functional hypothesis are discussed about the role played by olfaction in the economy of tongue muscle activity.

Antiepileptic treatment and oculomotor neurons.

This report describes the changes of the spontaneous firing rate due to an acute non-toxic dose of phenytoin (PHT), a drug commonly used in antiepileptic therapy, in the pre-motor neurons involved in saccadic movement. The drug (500 mg/kg of a 10% PHT suspension in arabic gum) was orally administered, and plasma and brain levels were regularly evaluated (EMIT assay). Results show that PHT significantly modifies the spontaneous electrical activity of the pre-motor neurons localized in the paramedian pontine reticular formation by inducing excitation, inhibition, or a biphasic effect. PHT action was observed 10-15 min after drug administration, when plasma and brain concentrations were still very low. The oculomotor system neurons appear to be a more specific target to this drug in comparison to the cerebellum and the vestibular system. Since the PHT action was observed 1 hour after drug administration in the vestibular nuclei and the cerebellum, which are extensively connected with the oculomotor neurons, it is possible to hypothesize that PHT can affect the oculomotor neurons directly and, with longer latency, indirectly through the vestibular nuclei and the cerebellum.

Brain electrobiogenesis protection induced by nimodipine and MK-801 during acute hypoxia in hypertensive rats.

The protective effect of nimodipine, a calcium entry blocker, and MK-801, an excitatory amino acid antagonist, on the cortical electrobiogenesis of hypertensive SHR-Charles-River adult rats during a 90-s session of acute hypoxia, was tested either with a single treatment or with a therapeutic association. Pretreatment with nimodipine alone (0.1 mg kg-1, i.v.) was particularly effective in increasing the rat brain resistence to hypoxia when compared to the controls. MK-801 pretreatment alone (3 mg kg-1, subcutaneous), improved significantly the recovery of brain electrical activity from hypoxia, as it not only shortened the recovery time but also abolished the post-hypoxic burst activity. The combined pretreatment with both drugs, using the same doses, displayed a more powerful brain protective effect on the cortical electrobiogenesis than the single drug pretreatment, suggesting the existence of a synergy between nimodipine and MK-801.

Effect of fetal hypoxia on seizure susceptibility in rats.

In the present study, susceptibility to Pentylentetrazol (PTZ)-induced seizures was tested in 45 four-wk old rats born to mothers exposed to moderate asphyxia in the last week of pregnancy by breathing N2 99.9% for 6 min in two separate sessions, (Group I--experimental rats) and in 44 rats of the same age, born after a normal pregnancy (Group II--controls). The results showed that the experimental rats, following episodes of asphyxia in intrauterine life, had a higher susceptibility to PTZ-induced seizures than the controls, manifested by earlier onset of convulsions and a higher incidence of fetal epileptic status. This occurred despite normal development and the absence of neurological deficits in the experimental rats in the first 4 wk of extrauterine life.