Horizontal projections of area 17 in Cebus monkeys: metric features, and modular and laminar distribution

Metric features and modular and laminar distributions of intrinsic projections of area 17 were studied in Cebus apella. Anterogradely and retrogradely labeled cell appendages were obtained using both saturated pellets and iontophoretic injections of biocytin into the operculum. Laminar and modular distributions of the labeled processes were analyzed using Nissl counterstaining, and/or cytochrome oxidase and/or NADPH-diaphorase histochemistry. We distinguished three labeled cell types: pyramidal, star pyramidal and stellate cells located in supragranular cortical layers (principally in layers IIIa, IIIb a, IIIb ß and IIIc). Three distinct axon terminal morphologies were found, i.e., Ia, Ib and II located in granular and supragranular layers. Both complete and partial segregation of group I axon terminals relative to the limits of the blobs of V1 were found. The results are compatible with recent evidence of incomplete segregation of visual information flow in V1 of Old and New World primates

Catabolism of Ap4A and Ap5A by rat brain synaptosomes

Adenosine 5',5'''-P1,P4-tetraphosphate (Ap4A) and adenosine 5',5'''-P1,P5-pentaphosphate (Ap5A) are stored in and released from rat brain synaptic terminals. In the present study we investigated the hydrolysis of dinucleotides (Ap4A and Ap5A) in synaptosomes from the cerebral cortex of adult rats. Ap4A and Ap5A, but not Ap3A, were hydrolyzed at pH 7.5 in the presence of 20 mM Tris/HCl, 2.0 mM MgCl2, 10 mM glucose and 225 mM sucrose at 37oC. The disappearance of the substrates measured by FPLC on a mono-Q HR column was both time and protein dependent. Since synaptosome integrity was at least 90% at the end of the assay, hydrolysis probably occurred by the action of an ecto-enzyme. Extracellular actions of adenine dinucleotides at central nervous system terminate due to the existence of ecto-nucleotidases which specifically cleave these dinucleotides. These enzymes in association with an ATP diphosphohydrolase and a 5'-nucleotidase are able to promote the complete hydrolysis of dinucleotides to adenosine in the synaptic cleft.

Melatonin modulation of presynaptic nicotinic acetylcholine receptors located on short noradrenergic neurons of the rat vas deferens: a pharmacological characterization

Melatonin, the pineal hormone produced during the dark phase of the light-dark cycle, modulates neuronal acetylcholine receptors located presynaptically on nerve terminals of the rat vas deferens. Recently we showed the presence of high affinity nicotine-binding sites during the light phase, and low and high affinity binding sites during the dark phase. The appearance of the low affinity binding sites was due to the nocturnal melatonin surge and could be mimicked by exposure to melatonin in vitro. The aim of the present research was to identify the receptor subtypes responsible for the functional response during the light and the dark phase. The rank order of potency of agonists was dimethylphenylpiperazinium (DMPP) = cytisine > nicotine > carbachol and DMPP = nicotine = cytisine > carbachol, during the light and dark phase, respectively, due to an increase in apparent affinity for nicotine. Mecamylamine similarly blocked the DMPP response during the light and the dark phase, while the response to nicotine was more efficiently blocked during the light phase. In contrast, methyllycaconitine inhibited the nicotine-induced response only at 21:00 h. Since a7 nicotinic acetylcholine receptors (nAChRs) have low affinity for nicotine in binding assays, we suggest that a mixed population composed of a3ß4 - plus a7-bearing nAChR subtypes is present at night. This plasticity in receptor subtypes is probably driven by melatonin since nicotine-induced contraction in organs from animals sacrificed at 15:00 h and incubated with melatonin (100 pg/ml, 4 h) is not totally blocked by mecamylamine. Thus melatonin, by acting directly on the short adrenergic neurons that innervate the rat vas deferens, induces the appearance of the low affinity binding site, probably an a7 nAChR subtype.

The role of nitric oxide in reproduction

Nitric oxide (NO) plays a crucial role in reproduction at every level in the organism. In the brain, it activates the release of luteinizing hormone-releasing hormone (LHRH). The axons of the LHRH neurons project to the mating centers in the brain stem and by afferent pathways evoke the lordosis reflex in female rats. In males, there is activation of NOergic terminals that release NO in the corpora cavernosa penis to induce erection by generation of cyclic guanosine monophosphate (cGMP). NO also activates the release of LHRH which reaches the pituitary and activates the release of gonadotropins by activating neural NO synthase (nNOS) in the pituitary gland. In the gonad, NO plays an important role in inducing ovulation and in causing luteolysis, whereas in the reproductive tract, it relaxes uterine muscle via cGMP and constricts it via prostaglandins (PG).

Selective destruction of nigrostriatal dopaminergic neurons does not alter [3H]-ryanodine binding in rat striatum

Dopamine nigrostriatal neurons are important for motor control and may contain a particularly dense population of ryanodine receptors involved in the control of dopamine release. To test this hypothesis, we used a classical model of unilateral selective lesion of these neurons in rats based on 6-hydroxydopamine (6-OHDA) injection into the substantia nigra. Binding of [3H]-GBR 12935, used as a presynaptic marker since it labels specifically the dopamine uptake complex, was dramatically decreased by 83-100% in striatum homogenates after 6-OHDA lesion. On the contrary, no reduction of [3H]-ryanodine binding was observed. The present data indicate that [3H]-ryanodine binding sites present in rat striatum are not preferentially localized in dopaminergic terminals.

Steps in the formation of neurites and synapses studied in cultured leech neurons

Leech neurons in culture have provided novel insights into the steps in the formation of neurite outgrowth patterns, target recognition and synapse formation. Identified adult neurons from the central nervous system of the leech can be removed individually and plated in culture under well-controlled conditions, where they retain their characteristic physiological properties, grow neurites and form specific chemical or electrical synapses. Different identified neurons develop distinctive outgrowth patterns that depend on their identities and on the molecular composition of the substrate. On native substrates, the patterns displayed by these neurons reproduce characteristics from the adult or the developing neurons. In addition, the substrate may induce selective directed growth between pairs of neurons that normally make contact in the ganglion. Upon contact, pairs of cultured leech neurons form chemical or electrical synapses, or both types depending on the neuronal identities. Anterograde and retrograde signals during membrane contact and synapse formation modify the distribution of synaptic terminals, calcium currents, and responses to 5-hydroxytryptamine.

Oxytocin is a cardiovascular hormone

Oxytocin (OT), a nonapeptide, was the first hormone to have its biological activities established and chemical structure determined. It was believed that OT is released from hypothalamic nerve terminals of the posterior hypophysis into the circulation where it stimulates uterine contractions during parturition, and milk ejection during lactation. However, equivalent concentrations of OT were found in the male hypophysis, and similar stimuli of OT release were determined for both sexes, suggesting other physiological functions. Indeed, recent studies indicate that OT is involved in cognition, tolerance, adaptation and complex sexual and maternal behaviour, as well as in the regulation of cardiovascular functions. It has long been known that OT induces natriuresis and causes a fall in mean arterial pressure, both after acute and chronic treatment, but the mechanism was not clear. The discovery of the natriuretic family shed new light on this matter. Atrial natriuretic peptide (ANP), a potent natriuretic and vasorelaxant hormone, originally isolated from rat atria, has been found at other sites, including the brain. Blood volume expansion causes ANP release that is believed to be important in the induction of natriuresis and diuresis, which in turn act to reduce the increase in blood volume. Neurohypophysectomy totally abolishes the ANP response to volume expansion. This indicates that one of the major hypophyseal peptides is responsible for ANP release. The role of ANP in OT-induced natriuresis was evaluated, and we hypothesized that the cardio-renal effects of OT are mediated by the release of ANP from the heart. To support this hypothesis, we have demonstrated the presence and synthesis of OT receptors in all heart compartments and the vasculature. The functionality of these receptors has been established by the ability of OT to induce ANP release from perfused heart or atrial slices. Furthermore, we have shown that the heart and large vessels like the aorta and vena cava are sites of OT synthesis. Therefore, locally produced OT may have important regulatory functions within the heart and vascular beds. Such functions may include slowing down of the heart or the regulation of local vascular tone.

Evidence of muscarinic acetylcholine receptors in the retinal centrifugal system of the chick

In this study we characterize the presence of muscarinic acetylcholine receptors (mAChR) in the isthmo-optic nucleus (ION) of chicks by immunohistochemistry with the M35 antibody. Some M35-immunoreactive fibers were observed emerging from the retinal optic nerve insertion, suggesting that they could be centrifugal fibers. Indeed, intraocular injections of cholera toxin B (CTb), a retrograde tracer, and double-labeling with M35 and CTb in the ION confirmed this hypothesis. The presence of M35-immunoreactive cells and the possible mAChR expression in ION and ectopic neuron cells in the chick brain strongly suggest the existence of such a cholinergic system in this nucleus and that acetylcholine release from amacrine cells may mediate interactions between retinal cells and ION terminals.

Aggrecan structure in amphibian cartilage

The structure of the large proteoglycan present in the bullfrog epiphyseal cartilage was studied by immunochemical and biochemical methods. The isolated monomer showed a polydisperse behavior on Sepharose CL2B, with a peak at Kav = 0.14. Chondroitin sulfate chains were identified by HPLC analysis of the products formed by chondroitinase digestion and mercuric acetate treatment. These chains have approximately 38 disaccharides, a Di45:Di68 ratio of 1.6 and GalNAc4S + GalNAc4,6S are the main non-reducing terminals. Keratan sulfate was identified by the use of two monoclonal antibodies in Western blots after chondroitinase ABC treatment. A keratan sulfate-rich region (~110 kDa) was isolated by sequential treatment with chondroitinase ABC and proteases. We also employed antibodies in Western blotting experiments and showed that the full length deglycosylated core protein is about 300 kDa after SDS-PAGE. Domain-specific antibodies revealed the presence of immunoreactive sites corresponding to G1/G2 and G3 globular domains and the characterization of this large proteoglycan as aggrecan. The results indicate the high conservation of the aggrecan domain structure in this lower vertebrate.

Effects of L-arginine on the diaphragm muscle twitches elicited at different frequencies of nerve stimulation

In rats, the nitric oxide (NO)-synthase pathway is present in skeletal muscle, vascular smooth muscle, and motor nerve terminals. Effects of NO were previously studied in rat neuromuscular preparations receiving low (0.2 Hz) or high (200 Hz) frequencies of stimulation. The latter frequency has always induced tetanic fade. However, in these previous studies we did not determine whether NO facilitates or impairs the neuromuscular transmission in preparations indirectly stimulated at frequencies which facilitate neuromuscular transmission. Thus, the present study was carried out to examine the effects of NO in rat neuromuscular preparations indirectly stimulated at 5 and 50 Hz. The amplitude of muscular contraction observed at the end (B) of a 10-s stimulation was taken as the ratio (R) of that obtained at the start (A) (R = B/A). S-nitroso-N-acetylpenicillamine (200 µM), superoxide dismutase (78 U/ml) and L-arginine (4.7 mM), but not D-arginine (4.7-9.4 mM), produced an increase in R (facilitation of neurotransmission) at 5 Hz. However, reduction in the R value (fade of transmission) was observed at 50 Hz. N G-nitro-L-arginine (8.0 mM) antagonized both the facilitatory and inhibitory effects of L-arginine (4.7 mM). The results suggest that NO may modulate the release of acetylcholine by motor nerve terminals.

Interhemispheric connections between primary visual areas: beyond the midline rule

In the last five years, a number of detailed anatomical, electrophysiological, optical imaging and simulation studies performed in a variety of non-human species have revealed that the functional organization of callosal connections between primary visual areas is more elaborate than previously thought. Callosal cell bodies and terminals are clustered in columns whose correspondence to features mapped in the visual cortex, such as orientation and ocularity, are starting to be understood. Callosal connections are not restricted to the vertical midline representation nor do they establish merely point-to-point retinotopic correspondences across the hemispheres, as traditionally believed. In addition, anatomical studies have revealed the existence of an ipsilateral component of callosal axons. The aim of this short review is to propose how these new data can be integrated into an updated scheme of the circuits responsible for assembling the primary visual field map.

Neo-Timm staining in the thalamus of chronically epileptic rats

The thalamus is an important modulator of seizures and is severely affected in cholinergic models of epilepsy. In the present study, chronically epileptic rats had their brains processed for neo-Timm and acetylcholinesterase two months after the induction of status epilepticus with pilocarpine. Both controls and pilocarpine-treated animals presented neo-Timm staining in the anterodorsal nucleus, laterodorsal nucleus, reticular nucleus, most intralaminar nuclei, nucleus reuniens, and rhomboid nucleus of the thalamus, as well as in the zona incerta. The intensity of neo-Timm staining was similar in control and pilocarpine-treated rats, except for the nucleus reuniens and the rhomboid nucleus, which had a lower intensity of staining in the epileptic group. In animal models of temporal lobe epilepsy, zinc seems to modulate glutamate release and to decrease seizure activity. In this context, a reduction of neo-Timm-stained terminals in the midline thalamus could ultimately result in an increased excitatory activity, not only within its related nuclei, but also in anatomical structures that receive their efferent connections. This might contribute to the pathological substrate observed in chronic pilocarpine-treated epileptic animals.

The inhibitory role of sympathetic nervous system in the Ca2+-dependent proteolysis of skeletal muscle

Mammalian cells contain several proteolytic systems to carry out the degradative processes and complex regulatory mechanisms to prevent excessive protein breakdown. Among these systems, the Ca2+-activated proteolytic system involves the cysteine proteases denoted calpains, and their inhibitor, calpastatin. Despite the rapid progress in molecular research on calpains and calpastatin, the physiological role and regulatory mechanisms of these proteins remain obscure. Interest in the adrenergic effect on Ca2+-dependent proteolysis has been stimulated by the finding that the administration of β2-agonists induces muscle hypertrophy and prevents the loss of muscle mass in a variety of pathologic conditions in which calpains are activated. This review summarizes evidence indicating that the sympathetic nervous system produces anabolic, protein-sparing effects on skeletal muscle protein metabolism. Studies are reviewed, which indicate that epinephrine secreted by the adrenal medulla and norepinephrine released from adrenergic terminals have inhibitory effects on Ca2+-dependent protein degradation, mainly in oxidative muscles, by increasing calpastatin levels. Evidence is also presented that this antiproteolytic effect, which occurs under both basal conditions and in stress situations, seems to be mediated by β2- and β3-adrenoceptors and cAMP-dependent pathways. The understanding of the precise mechanisms by which catecholamines promote muscle anabolic effects may have therapeutic value for the treatment of muscle-wasting conditions and may enhance muscle growth in farm species for economic and nutritional purposes.

The immunomodulator glatiramer acetate influences spinal motoneuron plasticity during the course of multiple sclerosis in an animal model

The immunomodulador glatiramer acetate (GA) has been shown to significantly reduce the severity of symptoms during the course of multiple sclerosis and in its animal model - experimental autoimmune encephalomyelitis (EAE). Since GA may influence the response of non-neuronal cells in the spinal cord, it is possible that, to some extent, this drug affects the synaptic changes induced during the exacerbation of EAE. In the present study, we investigated whether GA has a positive influence on the loss of inputs to the motoneurons during the course of EAE in rats. Lewis rats were subjected to EAE associated with GA or placebo treatment. The animals were sacrificed after 15 days of treatment and the spinal cords processed for immunohistochemical analysis and transmission electron microscopy. A correlation between the synaptic changes and glial activation was obtained by performing labeling of synaptophysin and glial fibrillary acidic protein using immunohistochemical analysis. Ultrastructural analysis of the terminals apposed to alpha motoneurons was also performed by electron transmission microscopy. Interestingly, although the GA treatment preserved synaptophysin labeling, it did not significantly reduce the glial reaction, indicating that inflammatory activity was still present. Also, ultrastructural analysis showed that GA treatment significantly prevented retraction of both F and S type terminals compared to placebo. The present results indicate that the immunomodulator GA has an influence on the stability of nerve terminals in the spinal cord, which in turn may contribute to its neuroprotective effects during the course of multiple sclerosis.

Sleep, ageing and night work

Studies have shown that the frequency or worsening of sleep disorders tends to increase with age and that the ability to perform circadian adjustments tends to decrease in individuals who work the night shift. This condition can cause consequences such as excessive sleepiness, which are often a factor in accidents that occur at work. The present study investigated the effects of age on the daytime and nighttime sleep patterns using polysomnography (PSG) of long-haul bus drivers working fixed night or day shifts. A total of 124 drivers, free of sleep disorders and grouped according to age (<45 years, N = 85, and ≥45 years, N = 39) and PSG timing (daytime (D) PSG, N = 60; nighttime (N) PSG, N = 64) participated in the study. We observed a significant effect of bedtime (D vs N) and found that the length of daytime sleep was shorter [D: <45 years (336.10 ± 73.75 min) vs N: <45 years (398 ± 78.79 min) and D: ≥45 years (346.57 ± 43.17 min) vs N: ≥45 years (386.44 ± 52.92 min); P ≤ 0.05]. Daytime sleep was less efficient compared to nighttime sleep [D: <45 years (78.86 ± 13.30%) vs N: <45 years (86.45 ± 9.77%) and D: ≥45 years (79.89 ± 9.45%) and N: ≥45 years (83.13 ± 9.13%); P ≤ 0.05]. An effect of age was observed for rapid eye movement sleep [D: <45 years (18.05 ± 6.12%) vs D: ≥45 years (15.48 ± 7.11%) and N: <45 years (23.88 ± 6.75%) vs N: ≥45 years (20.77 ± 5.64%); P ≤ 0.05], which was greater in younger drivers. These findings are inconsistent with the notion that older night workers are more adversely affected than younger night workers by the challenge of attempting to rest during the day.