The Intrahippocampal Infusion Of Crotamine From Crotalus Durissus Terrificus Venom Enhances Memory Persistence In Rats.

Previous research has shown that crotamine, a toxin isolated from the venom of Crotalus durissus terrificus, induces the release of acetylcholine and dopamine in the central nervous system of rats. Particularly, these neurotransmitters are important modulators of memory processes. Therefore, in this study we investigated the effects of crotamine infusion on persistence of memory in rats. We verified that the intrahippocampal infusion of crotamine (1 μg/μl; 1 μl/side) improved the persistence of object recognition and aversive memory. By other side, the intrahippocampal infusion of the toxin did not alter locomotor and exploratory activities, anxiety or pain threshold. These results demonstrate a future prospect of using crotamine as potential pharmacological tool to treat diseases involving memory impairment, although it is still necessary more researches to better elucidate the crotamine effects on hippocampus and memory.

Learning processes and the neural analysis of conditioning

Classical and operant conditioning principles, such as the behavioral discrepancy-derived assumption that reinforcement always selects antecedent stimulus and response relations, have been studied at the neural level, mainly by observing the strengthening of neuronal responses or synaptic connections. A review of the literature on the neural basis of behavior provided extensive scientific data that indicate a synthesis between the two conditioning processes based mainly on stimulus control in learning tasks. The resulting analysis revealed the following aspects. Dopamine acts as a behavioral discrepancy signal in the midbrain pathway of positive reinforcement, leading toward the nucleus accumbens. Dopamine modulates both types of conditioning in the Aplysia mollusk and in mammals. In vivo and in vitro mollusk preparations show convergence of both types of conditioning in the same motor neuron. Frontal cortical neurons are involved in behavioral discrimination in reversal and extinction procedures, and these neurons preferentially deliver glutamate through conditioned stimulus or discriminative stimulus pathways. Discriminative neural responses can reliably precede operant movements and can also be common to stimuli that share complex symbolic relations. The present article discusses convergent and divergent points between conditioning paradigms at the neural level of analysis to advance our knowledge on reinforcement.

Effects of ascorbic acid supplementation in ileum myenteric neurons of streptozotocin-induced diabetic rats

The exacerbation of the oxidative stress and of the polyol pathway which impair damage myenteric plexus are metabolic characteristics of diabetes. The ascorbic acid (AA) is an antioxidant and an aldose reductase inhibitor, which may act as neuroprotector. The effects of AA supplementation on the density and cellular body profile area (CP) of myenteric neurons in STZ-induced diabetes in rats were assessed. Four groups with five animals each were formed: normoglycemic (C); diabetic (D); AA-treated diabetic (DS) and AA-treated normoglycemic (CS). Dosagen of 50mg of AA were given, three times a week, for each animal (group DS and CS). Ninety days later and after euthanasia, the ileum was collected and processed for the NADPH-diaphorase technique. There were no differences (P>0.05) in the neuronal density among the groups. The CP area was lower (P<0.05) in the DS and CS groups, with a higher incidence of neurons with a CP area exceeding 200µm² for groups C and D. The AA had no influence on the neuronal density in the ileum but had a neuroprotective effect, preventing the increase in the CP area and allowing a higher number of neurons with a CP area with less than 200µm².

Dopamine-beta hydroxylase polymorphism and cocaine addiction

Cocaine addiction involves a number of medical, psychological and social problems. Understanding the genetic aetiology of this disorder will be essential for design of effective treatments. Dopamine-beta hydroxylase (DbH) catalyzes the conversion of dopamine to norepinephrine and could, therefore, have an influence on both cocaine action and the basal sensitivity of neurotransmitter systems to cocaine. Recently, the - 1021C> T polymorphism have been found to strongly correlated with individual variation in plasma DbH activity. To test the influence of this polymorphism on the susceptibility of cocaine addiction, we decided to genotype it in a sample of 689 cocaine addicts and 832 healthy individuals. Genotypic and allelic analyses did not show any evidence of association with cocaine addiction, even after correcting for the effect of population stratification and other possible confounders. Our results do not support a major role of the - 1021C> T polymorphism or the gene itself in the development of cocaine addiction but further examination of other variants within this gene will be necessary to completely rule out an effect.

Actions of angiotensin II and dopamine in the medial preoptic area on prolactin secretion

Dopamine (DA) is known as a primary regulator of prolactin secretion (PRL) and angiotensin II (Ang II) has been recognized as one brain inhibitory factor of this secretion. In this work, estrogen-primed or unprimed ovariectornized rats were submitted to the microinjection of saline or Ang II after previous microinjection of saline or of DA antagonist (haloperidol, sulpiride or SCH) both in the medial preoptic area (MPOA). Our study of these interactions has shown that 1) estrogen-induced PRL secretion is mediated by Ang II and DA actions in the MPOA, i.e. very high plasma PRL would be prevented by inhibitory action of Ang II, while very low levels would be prevented in part by stimulatory action of DA through D-2 receptors, 2) the inhibitory action of Ang II depends on estrogen and is mediated in part by inhibitory action of DA through D, receptors and in other part by inhibition of stimulatory action of DA through D2 receptors.

Effects of protein deprivation and re-feeding on P2X(2) receptors in enteric neurons

AIM: To investigate the effects of malnutrition and refeeding on the P2X(2) receptor, nitric oxide synthase (NOS), calretinin, calbindin and choline acetyltransferase (ChAT) in neurons of the rat ileum. METHODS: We analyzed the co-localization, numbers and sizes of P2X(2)-expressing neurons in relation to NOS-IR (immunoreactive), calbindin-IR, ChAT-IR, and calretinin-IR neurons of the myenteric and submucosal plexus. The experimental groups consisted of: (1) rats maintained on normal feed throughout pregnancy until 42 d post-parturition (N); (2) rats deprived of protein throughout pregnancy and 42 d post-parturition (D); and (3) rats undernourished for 21 d post-parturition and then given a protein diet from days 22 to 42 (DR). The myenteric and submucosal plexuses were evaluated by double labeling by immunohistochemical methods for P2X(2) receptor, NOS, ChAT, calbindin and calretinin. RESULTS: We found similar P2X(2) receptor immunoreactivity in the cytoplasm and surface membranes of myenteric and submucosal neurons from the N, D and DR groups. Double labeling of the myenteric plexus demonstrated that approximately 100% of NOS-IR, calbindin-IR, calretinin-IR and ChAT-IR neurons in all groups also expressed the P2X(2) receptor. In the submucosal plexus, the calretinin-IR, ChAT-IR and calbindinIR neurons were nearly all immunoreactive for the P2X(2) receptor. In the myenteric plexus, there was a 19% increase in numbers per cm(2) for P2X(2) receptor-IR neurons, 64% for NOS-IR, 84% for calretinin-IR and 26% for ChAT-IR neurons in the D group. The spatial density of calbindin-IR neurons, however, did not differ among the three groups. The submucosal neuronal density increased for calbindin-IR, calretinin-IR and ChAT-IR neurons. The average size of neurons in the myenteric plexus neurons in the D group was less than that in the controls and, in the re-fed rats; there was a 34% reduction in size only for the calretinin-IR neurons. CONCLUSION: This work demonstrates that expression of the P2X(2) receptor is present in inhibitory, intrinsic primary afferent, cholinergic secretomotor and vasomotor neurons. Undernutrition affected P2X(2) receptor expression in the submucosal plexus, and neuronal and size. These changes were rescued in the re-fed rats. (C) 2010 Baishideng. All rights reserved.

Leptin's effect on puberty in mice is relayed by the ventral premammillary nucleus and does not require signaling in Kiss1 neurons

Studies m hum ins and rodents indicate that a minimum amount of stored energy is required for normal pubertal development The adipocyte-derived hormone leptin is a key metabolic signal to the neuroendocrine reproductive axis Humans and mice lacking leptin or the leptin receptor (LepR) (ob/ob and db/db mice, respectively) are infertile and fail to enter puberty Leptin administration to leptin-deficient subjects and ob/ob mice induces puberty and restores fertility, but the exact site or sites of leptin action are unclear Here, we found that genetic deletion of LepR selectively from hypothalamic Kiss1 neurons m mice had no effect on puberty or fertility, indicating that direct leptin signaling m Kiss1 neurons is not required for these processes However, bilateral lesions of the ventral premammillary nucleus (PMV) of ob/ob mice blunted the ability of exogenous leptin to induce sexual maturation Moreover, unilateral reexpression of endogenous LepR m PMV neurons was sufficient to induce puberty and improve fertility m female LepR-null mice This LepR reexpression also normalized the increased hypothalamic GnRH content characteristic of leptin-signaling deficiency These data suggest that the PMV is a key site for leptin's permissive action at the onset of puberty and support the hypothesis that the multiple actions of leptin to control metabolism and reproduction at e anatomically dissociated

Connectivity and dynamics of neuronal networks as defined by the shape of individual neurons

Biological neuronal networks constitute a special class of dynamical systems, as they are formed by individual geometrical components, namely the neurons. In the existing literature, relatively little attention has been given to the influence of neuron shape on the overall connectivity and dynamics of the emerging networks. The current work addresses this issue by considering simplified neuronal shapes consisting of circular regions (soma/axons) with spokes (dendrites). Networks are grown by placing these patterns randomly in the two-dimensional (2D) plane and establishing connections whenever a piece of dendrite falls inside an axon. Several topological and dynamical properties of the resulting graph are measured, including the degree distribution, clustering coefficients, symmetry of connections, size of the largest connected component, as well as three hierarchical measurements of the local topology. By varying the number of processes of the individual basic patterns, we can quantify relationships between the individual neuronal shape and the topological and dynamical features of the networks. Integrate-and-fire dynamics on these networks is also investigated with respect to transient activation from a source node, indicating that long-range connections play an important role in the propagation of avalanches.

Fluorescent indication that nitric oxide formation in NTS neurons is modulated by glutamate and GABA

Nitric oxide (NO) in NTS plays an important role in regulating autonomic function to the cardiovascular system. Using the fluorescent dye DAF-2 DA, we evaluated the NO concentration in NTS. Brainstem slices of rats were loaded with DAF-2 DA, washed, fixed in paraformaldehyde and examined under fluorescent light. In different experimental groups, NTS slices were pre-incubated with 1 mM L-NAME (a non-selective NOS inhibitor), 1 MM D-NAME (an inactive enantiomere of L-NAME), 1 mM kynurenic acid (a nonselective ionotropic receptors antagonist) or 20 mu M bicuculline (a selective GABA(A) receptors antagonist) before and during DAF-2 DA loading. Images were acquired using a confocal microscope and the intensity of fluorescence was quantified in three antero-posterior NTS regions. In addition, slices previously loaded with DAF-2 DA were incubated with NeuN or GFAP antibody. A semi-quantitative analysis of the fluorescence intensity showed that the basal NO concentration was similar in all antero-posterior aspects of the NTS (rostral intermediate, 15.5 +/- 0.8 AU: caudal intermediate, 13.2 +/- 1.4 AU; caudal commissural, 13.8 +/- 1.4 AU, n = 10). In addition, the inhibition of NOS and the antagonism of glutamatergic receptors decreased the NO fluorescence in the NTS. On the other hand, D-NAME did not affect the NO fluorescence and the antagonism of GABAA receptors increased the NO fluorescence in the NTS. It is important to note that the fluorescence for NO was detected mainly in neurons. These data show that the fluorescence observed after NTS loading with DAF-2 DA is a result of NO present in the NTS and support the concept that NTS neurons have basal NO production which is modulated by L-glutamate and GABA. (C) 2009 Elsevier Inc. All rights reserved.

Characterization of calcium-activated chloride channels in patches excised from the dendritic knob of mammalian olfactory receptor neurons

We investigated the properties of calcium-activated chloride channels in inside-out membrane patches from the dendritic knobs of acutely dissociated rat olfactory receptor neurons. Patches typically contained large calcium-activated currents, with total conductances in the range 30-75 nS. The dose response curve for calcium exhibited an EC50 of about 26 mu M. In symmetrical NaCl solutions, the current-voltage relationship reversed at 0 mV and was linear between -80 and +70 mV. When the intracellular NaCl concentration was progressively reduced from 150 to 25 mM, the reversal potential changed in a manner consistent with a chloride-selective conductance. Indeed, modeling these data with the Goldman-Hodgkin-Katz equation revealed a P-Na/P-Cl of 0.034. The halide permeability sequence was P-Cl > P-F > P-I > P-Br indicating that permeation through the channel was dominated by ion binding sites with a high field strength. The channels were also permeable to the large organic anions, SCN-, acetate(-), and gluconate(-), with the permeability sequence P-Cl > P-SCN > gluconaie. Significant permeation to gluconate ions suggested that the channel pore had a minimum diameter of at least 5.8 Angstrom.

Hypothalamic dopamine D1 receptors are involved in the stimulation of prolactin secretion by high environmental temperature in the female sheep

Recent evidence suggests that dopamine, acting via its D1 receptors, may function as a neurotransmitter in intrahypothalamic pathways involved in the stimulation of prolactin secretion. Functional dopamine D1 receptors are present in the ventromedial hypothalamic nucleus (VMH) and we hypothesized that they might be part of a prolactin-stimulatory pathway activated by stress. We tested this hypothesis in a series of experiments on sheep involving two different forms of stressors, audiovisual (barking dog) and high environmental temperature. We attempted to block the stimulation of prolactin secretion by infusion into the VMH of an antagonist specific for the D1 receptor. Ovariectomised, oestradiol-implanted merino ewes were surgically implanted with bilateral guide tubes directed at the VMH. After a 180 min pretreatment period, the ewes either were or were not exposed to a stressor (30 min of barking dog or 120 min at 35 degrees C, 65% relative humidity). D1 receptor antagonist, SCH23390 or vehicle (0.9% saline) was infused into the VMH (1.7 mu l/h, 120 nmol/h) for 60 min prior to and during the stressor period. Blood was sampled every 15 min via jugular cannulae and the plasma was assayed for prolactin, cortisol and growth hormone (GH). Both stressors significantly increased prolactin concentrations over control levels. SCH23390 infusion significantly attenuated the prolactin response to high environmental temperature, but had no effect on the prolactin response to audiovisual stress. Cortisol concentrations were significantly increased by audiovisual stress only and were not affected by SCH23390, GH concentrations were not changed by either stressor or infusion. Drug infusion alone did not affect the concentration of the hormones. The data suggest that the VMH D1 receptors are involved in a prolactin stimulatory pathway in response to high environmental temperature. The inability of the D1 antagonist to affect the response to the barking dog indicates that this pathway is stress-specific, implying that there is more than one mechanism or pathway involved in the prolactin response to different stressors.

New techniques for biopsy and culture of human olfactory epithelial neurons

Objective: To improve the success of culturing olfactory neurons from human nasal mucosa by investigating the intranasal distribution of the olfactory epithelium and devising new techniques for growing human olfactory epithelium in vitro. Design: Ninety-seven biopsy specimens were obtained from 33 individuals, aged 21 to 74 years, collected from 6 regions of the nasal cavity. Each biopsy specimen was bisected, and 1 piece was processed for immunohistochemistry or electron microscopy while the other piece was dissected further for explant culture. Four culture techniques were performed, including whole explants and explanted biopsy slices. Five days after plating, neuronal differentiation was induced by means of a medium that contained basic fibroblast growth factor. After another 5 days, cultures were processed for immunocytochemical analysis. Results: The probability of finding olfactory epithelium in a biopsy specimen ranged from 30% to 76%, depending on its location. The dorsoposterior regions of the nasal septum and the superior turbinate provided the highest probability, but, surprisingly, olfactory epithelium was also found anteriorly and ventrally on both septum and turbinates. A new method of culturing the olfactory epithelium was devised. This slice culture technique improved the success rate for generating olfactory neurons from 10% to 90%. Conclusions: This study explains and overcomes most of the variability in the success in observing neurogenesis in cultures of adult human olfactory epithelium. The techniques presented here make the human olfactory epithelium a useful model for clinical research into certain olfactory dysfunctions and a model for the causes of neurodevelopmental and neurodegenerative diseases.

Excitatory synaptic inputs to pyramidal neurons of the lateral amygdala

Whole-cell patch clamp recordings were made from pyramidal neurons in the rat lateral amygdala (LA). Synaptic currents were evoked by stimulating in either the external capsule (ec), internal capsule (ic) or basolateral nucleus (BLA). Stimulation of either the ic, ec or BLA evoked a glutamatergic excitatory synaptic current (EPSC) which was mediated by both non-NMDA and NMDA (N-methyl-D-aspartic acid) receptors, The ratio of the amplitude of the NMDA receptor-mediated component measured at +40 mV to the amplitude of the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) component measured at -60 mV was similar regardless of whether EPSCs were evoked in the ec, ic or BLA. At resting membrane potentials, excitatory synaptic potentials evoked from either the ec or putative thalamic inputs were unaffected by application of the NMDA receptor antagonist APV. Spontaneous glutamatergic currents had two components to their decay phase. The slow component was selectively blocked by the NMDA receptor antagonist D-APV, indicating that AMPA and NMDA receptors are colocalized in spiny neurons. We conclude that pyramidal cells of the LA receive convergent inputs from the cortex, thalamus and basal nuclei. At all inputs, both AMPA/kainate and NMDA-type receptors are active and colocalized in the postsynaptic density.

Photolytic manipulation of [Ca2+](i) reveals slow kinetics of potassium channels underlying the afterhyperpolarization in hipppocampal pyramidal neurons

The identity of the potassium channel underlying the slow, apamin-insensitive component of the afterhyperpolarization current (sl(AHP)) remains unknown. We studied sl(AHP) in CA1 pyramidal neurons using simultaneous whole-cell recording, calcium fluorescence imaging, and flash photolysis of caged compounds. Intracellular calcium concentration ([Ca2+](i)) peaked earlier and decayed more rapidly than sl(AHP). Loading cells with low concentrations of the calcium chelator EGTA slowed the activation and decay of sl(AHP). In the presence of EGTA, intracellular calcium decayed with two time constants. When [Ca2+](i) was increased rapidly after photolysis of DM-Nitrophen, both apamin-sensitive and apamin-insensitive outward currents were activated. The apamin-sensitive current activated rapidly (<20 msec), whereas the apamin-insensitive current activated more slowly (180 msec). The apamin-insensitive current was reduced by application of serotonin and carbachol, confirming that it was caused by sl(AHP) channels. When [Ca2+](i) was decreased rapidly via photolysis of diazo-2, the decay of sl(AHP) was similar to control (1.7 sec). All results could be reproduced by a model potassium channel gated by calcium, suggesting that the channels underlying sl(AHP) have intrinsically slow kinetics because of their high affinity for calcium.

Neurons in the hilus region of the rat hippocampus are depleted in number by exposure to alcohol during early postnatal life

We have previously shown that exposing rats to a relatively high dose of ethanol during early postnatal life resulted in a deficit in spatial learning ability. This ability is controlled, at least in part, by the hippocampal formation. The purpose of the present study was to determine whether exposure of rats to ethanol during early postnatal life affected the number of specific neurons in the hippocampus. Wistar rats were exposed to a relatively high daily dose of ethanol between postnatal days 10 and 15 by placing them for 3 h each day in a chamber containing ethanol vapor. The blood ethanol concentration was about 430 mg/dl at the end of the exposure period. Groups of ethanol-treated (ET) rats, separation controls (SC), and mother-reared controls (MRC) were anesthetized and killed at 16 days of age by perfusion with phosphate-buffered glutaraldehyde (2.5%). The Cavalieri principle was used to determine the volume of various subdivisions of the hippocampal formation (CA1, CA2+CA3, hilus, and granule cell layer), and the physical disector method was used to estimate the numerical densities of neurons within each subdivision. The total number of neurons was calculated by multiplying estimates of the numerical density with the volume. There were, on average, about 441,000 granule cells in the granule cell layer and 153,000 to 177,000 pyramidal cells in both the CA1 and CA2+CA3 regions in all three treatment groups. In the hilus region, ET rats had about 27,000 neuronal cells. This was significantly fewer than the average of 38,000 such neurons estimated to be present in both MRC and SC animals. Thus, neurons in the hilus region may be particularly vulnerable to the effects of a high dose of ethanol exposure during early postnatal life. (C) 2000 Wiley-Liss, Inc.