Effect of hypoxia in early perinatal ontogenesis on behavior and structural characteristics of the rat brain

The study has shown that acute hypoxia in newborn rat pups can lead to disturbances in processes of formation of brain structures, behavior reactions, and learning in the subsequent ontogenesis. The single normobaric hypoxia at the 2nd day of life causes retardation of such integrative parameter or general development and growth as body mass at the period of feeding. In such animals, essential disturbances of the sensorimotor development were revealed in forms of delay of reflex reactions of turning on a plane, negative geotaxis, and avoidance of edge. Also detected was action of hypoxia on hanging on a rope by using front legs (a symptom of muscle weakness). Morphological study has shown all studied functional zones of neocortex (motor, sensomotor, auditory, visual) to response stereotypically to the early postnatal action of hypoxia. The death of nerve cells is predominant in the II–III associative layers, the sizes and number of pyramidal neurons are sharply decreased. Different hippocampus fields maturing after death in mammals show a characteristic response to hypoxia. In individual hippocampus fields there was detected a different degree of death of neurons, with predominance in the CA3 and CA4 fields. A possibility of modeling the perinatal encephalopathy with minimal brain dysfunctions in children is discussed.     

GABA itself promotes the developmental switch of neuronal GABAergic responses from excitation to inhibition

GABA is the main inhibitory neurotransmitter in the adult brain. Early in development, however, GABAergic synaptic transmission is excitatory and can exert widespread trophic effects. During the postnatal period, GABAergic responses undergo a switch from being excitatory to inhibitory. Here, we show that the switch is delayed by chronic blockade of GABA(A) receptors, and accelerated by increased GABA(A) receptor activation. In contrast, blockade of glutamatergic transmission or action potentials has no effect. Furthermore, GABAergic activity modulated the mRNA levels of KCC2, a K(+)-Cl(-) cotransporter whose expression correlates with the switch. Finally, we report that GABA can alter the properties of depolarization-induced Ca(2+) influx. Thus, GABA acts as a self-limiting trophic factor during neural development.     

Formation of GABA-Aα1 and GABA-B1 receptor-mediated inhibitory network in the ventrolateral part of the solitary tract nucleus during the early postnatal period under normal conditions and prenatal serotonin deficiency

Time course of formation of the inhibitory receptor network (GABA-Aα1 and GABA-B1) in the respiratory subnuclei (ventral and lateral) of the solitary tract (NTS) during the early postnatal period was studied on laboratory Wistar rats under normal conditions and prenatal serotonin deficiency. It was found that in normal rats the maturation of the inhibitory receptor networks in both NTS subnuclei occurs within the first three postnatal weeks. Some features of their formation were noticed. The dynamics of changes in intensity of GABA-Аα1 expression in the ventral and lateral subnuclei proceeds in a similar way. During the first postnatal week, GABA-Aα1 expression is low. In the neuropil, the network of GABA-Aα1-containing presynaptic terminals and synapses is poorly developed. Within the second week, the number of GABA-Aα1-expressing neurons increases in both subnuclei with a simultaneous rise in the density of the network of terminals and synaptic structures. By the end of the third week, the number of GABA-Aα1-expressing neurons decreases, but the network density continues to increase. GABA-B1 expression in the ventral and lateral subnuclei occurs also simultaneously, although with some distinctions. During the first postnatal week, intensity of GABA-B1 expression is weak. In the neuropil, few GABA-B1-containing terminals form a loose network with sporadic synaptic structures. During the second week, expression of the receptor increases, being particularly considerable in the ventral subnuclei. Simultaneously, the density of the presynaptic terminals increases. By the end of the third week, the number of GABA-B1-expressing neurons in the ventral subnuclei decreases, while in the lateral subnuclei remains almost intact and the network density increases. The data obtained show that prenatal serotonin deficiency leads to malformation and impaired maturation of the GABAergic inhibitory neuronal network in the ventrolateral part of NTS.     

Effect of retinal ablation on the expression of calbindin D28k and GABA in the chick optic tectum

The effect of retinal ablation on qualitative and quantitative changes of calbindin D28k and GABA expression in the contralateral optic tectum was studied in young chicks. Fifteen days old chicks had unilateral retinal ablation and after 7 or 15 days, calbindin expression was analyzed by Western blot and immunocytochemistry. Neuronal degeneration was followed by the amino-cupric silver technique. After 15 days, retinal lesions produced a significant decrease in calbindin immunostaining in the neuropil of layers 5-6 and in the somata of neurons from the layers 8 and 10 of the contralateral tectum, being this effect less marked at 7 days post-lesion. Double staining revealed that 50-60% of cells in the layers 8 and 10 were calbindin and GABA positive, 30-45% were only calbindin positive and 5-10% were only GABAergic neurons. Retinal ablation also produced a decrease in the GABA expression at either 7 or 15 days after surgery. At 7 days, dense silver staining was observed in the layers 5-6 from the optic tectum contralateral to the retinal ablation, which mainly represented neuropil that would come from processes of retinal ganglion cells. Tectal neuronal bodies were not stained with silver, although some neurons were surrounded by coarse granular silver deposits. In conclusion, most of calbindin molecules are present in neurons of the tectal GABAergic inhibitory circuitry, whose functioning apparently depends on the integrity of the visual input. A possible role of calbindin in the control of intracellular Ca2+ in neurons of this circuit when the visual transmission arrives to the optic tectum remains to be studied.     

Comparative studies on the GABA-transaminase immunoreactivity in rat and gerbil brains.

Gamma-aminobutyric acid (GABA) is the most important inhibitory neurotransmitter in the central nervous system (CNS). Degradation of GABA in the CNS is catalyzed by the action of GABA transaminase (GABA-T). However, the neuroanatomical characteristics of GABA-T in the gerbil, which is a useful experimental animal in neuroscience, are still unknown. Therefore, we performed a comparative analysis of the distribution of GABA-T in rat and gerbil brains using immunohistochemistry. GABA-T immunoreactive neurons were observed in the regions which contained GABAergic neurons of both animals: corpus striatum; substantia nigra, pars reticulata; septal nucleus; and accumbens nucleus. GABA-T + neurons were restricted to layers III and V in the rat. Unlike the rat GABA-T + neurons were observed in layers II, III, and V of the gerbil cerebral cortex. These results suggest that the expression of GABA-T in the gerbil brain may be similar to that in the rat brain, except in the cerebral cortex.     

Non-associative Potentiation of Perisomatic Inhibition Alters the Temporal Coding of Neocortical Layer 5 Pyramidal Neurons

In the neocortex, the coexistence of temporally locked excitation and inhibition governs complex network activity underlying cognitive functions, and is believed to be altered in several brain diseases. Here we show that this equilibrium can be unlocked by increased activity of layer 5 pyramidal neurons of the mouse neocortex. Somatic depolarization or short bursts of action potentials of layer 5 pyramidal neurons induced a selective long-term potentiation of GABAergic synapses (LTPi) without affecting glutamatergic inputs. Remarkably, LTPi was selective for perisomatic inhibition from parvalbumin basket cells, leaving dendritic inhibition intact. It relied on retrograde signaling of nitric oxide, which persistently altered presynaptic GABA release and diffused to inhibitory synapses impinging on adjacent pyramidal neurons. LTPi reduced the time window of synaptic summation and increased the temporal precision of spike generation. Thus, increases in single cortical pyramidal neuron activity can induce an interneuron-selective GABAergic plasticity effectively altering the computation of temporally coded information.     

Ultrastructure of the serotoninergic system of the motor area of the cerebral cortex

Owing to the microscopical investigation, using selective neurotoxin 5,7-dihydroxytryptamine, it has been possible to reveal the serotoninergic system and targets of its innervation in the rat cerebral cortex motor area. The serotoninergic axonal varicosities and synaptic boutons are present in all layers of the neocortex. Their large amount is revealed in the I and II layers. The terminals form contacts with dendrites of small size, sometimes they terminate on the head of the spines, as well as on bodies of neurons in different layers. According to their position and ultrastructural organization these neurons are, perhaps, pyramidal, that is glutamatergic, and those less in their size--refer to interneurons and can be GABAergic ones. Basing on own data and those of the literature, concerning the existence of nonsynaptic link for transmission of serotoninergic effects, a conclusion is made that a coordinating functioning of the synaptic and non-synaptic intercellular integrative mechanisms ensure a wide range of functions of the serotoninergic system in the cerebral cortex.     

GABAergic interneuron to astrocyte signalling: a neglected form of cell communication in the brain

GABAergic interneurons represent a minority of all cortical neurons and yet they efficiently control neural network activities in all brain areas. In parallel, glial cell astrocytes exert a broad control of brain tissue homeostasis and metabolism, modulate synaptic transmission and contribute to brain information processing in a dynamic interaction with neurons that is finely regulated in time and space. As most studies have focused on glutamatergic neurons and excitatory transmission, our knowledge of functional interactions between GABAergic interneurons and astrocytes is largely defective. Here, we critically discuss the currently available literature that hints at a potential relevance of this specific signalling in brain function. Astrocytes can respond to GABA through different mechanisms that include GABA receptors and transporters. GABA-activated astrocytes can, in turn, modulate local neuronal activity by releasing gliotransmitters including glutamate and ATP. In addition, astrocyte activation by different signals can modulate GABAergic neurotransmission. Full clarification of the reciprocal signalling between different GABAergic interneurons and astrocytes will improve our understanding of brain network complexity and has the potential to unveil novel therapeutic strategies for brain disorders.     

A light and electron microscopic study of betaine/GABA transporter distribution in the monkey cerebral neocortex and hippocampus

The present study aimed to elucidate the distribution of betaine/γ-aminobutyric acid (GABA) transporter-1 (BGT-1) in the normal monkey cerebral neocortex and hippocampus by immunoperoxidase and Immunogold labelling. BGT-1 was observed in pyramidal neurons in the cerebral neocortex and the CA fields of the hippocampus. Large numbers of small diameter dendrites or dendritic spines were observed in the neuropil. These made asymmetrical synaptic contacts with unlabelled axon terminals containing small round vesicles, characteristic of glutamatergic terminals. BGT-1 label was observed in an extra-perisynaptic region, away from the post-synaptic density. Immunoreactivity was not observed in portions of dendrites that formed symmetrical synapses, axon terminals, or glial cells. The distribution of BGT-1 on dendritic spines, rather than at GABAergic axon terminals, suggests that the transporter is unlikely to play a major role in terminating the action of GABA at a synapse. Instead, the osmolyte betaine is more likely to be the physiological substrate of BGT-1 in the brain, and the presence of the transporter in pyramidal neurons suggests that these neurons utilize betaine to maintain osmolarity.     

Postnatal maturation of gamma-aminobutyric acidA and B-mediated inhibition in the CA3 hippocampal region of the rat

In the adult central nervous system, GABAergic synaptic inhibition is known to play a crucial role in preventing the spread of excitatory glutamatergic activity. This inhibition is achieved by a membrane hyperpolarization through the activation of postsynaptic γ-aminobutyric acid_A (GABA_A) and GABA_B receptors. In addition, GABA also depress transmitter release acting through presynaptic GABA_B receptors. Despite the wealth of data regarding the role of GABA in regulating the degree of synchronous activity in the adult, little is known about GABA transmission during early stages of development. In the following we report that GABA mediates most of the excitatory drive at early stages of development in the hippocampal CA3 region. Activation of GABA_A receptors induces a depolarization and excitation of immature CA3 pyramidal neurons and increases intracellular Ca²⁺ ([Ca²⁺]_i) during the first postnatal week of life. During the same developmental period, the postsynaptic GABA_B-mediated inhibition is poorly developed. In contrast, the presynaptic GABA_B-mediated inhibition is well developed at birth and plays a crucial role in modulating the postsynaptic activity by depressing transmitter release at early postnatal stages. We have also shown that GABA plays a trophic role in the neuritic outgrowth of cultured hippocampal neurons. © 1995 John Wiley & Sons, Inc.     

The remote consequences of hypoxia influence in perinatal development period on structurally functional characteristics of the rat brain

The study has shown that influence of acute hypoxia in perinatal period leads to structural changes in motor and visual of the neocortex for 20 postnatal days in the form of disturbance of the structural organisation of the neocortex layers. Different fields of hippocampus in perinatal period differently react to hypoxia, and evidence of existence of a long-term perinatal hypoxia was obtained. It is established that after action of acute hypoxia in all the fields there is a cellular destruction, and thinning of pyramidal neurones layers. The most expressed cellular destruction takes place in fields CA4 and CA3. In process of augmentation, the destruction of cells remains appreciable in the field CA4, reduced in the field CA3 and not found in the field CA1; however, in fascia dentate, the destruction of granular neurones with age augmentation increases. Along with in reduction of the dimensions of cellular bodies pyramidal neurones in all fields of hippocampus takes place. Also in all fields of hippocampus, activation of astrocytic reaction occurs, more expressed in the field CA4. The hypoxia influence in the early postnatal period can affect synaptogenes, particularly the formation of giant synapses in a dentate fascia. Study of functional features of the excitatory system of such animals has shown that hypoxia can induce appreciable disturbances of behavioural responses. In rats, disturbances of inhibiting functions of the cerebral cortex, raised anxiety, and spatial learning and working memory disturbances have been noted.     

Developmental change and sexual difference in synaptic modulation produced by oxytocin in rat substantia gelatinosa neurons

We have previously reported that oxytocin produces an inward current at a holding potential of ?70 mV without a change in glutamatergic excitatory transmission in adult male rat spinal lamina II (substantia gelatinosa; SG) neurons that play a pivotal role in regulating nociceptive transmission. Oxytocin also enhanced GABAergic and glycinergic spontaneous inhibitory transmissions in a manner sensitive to a voltage-gated Na⁺-channel blocker tetrodotoxin. These actions were mediated by oxytocin-receptor activation. Such a result was different from that obtained by other investigators in young male rat superficial dorsal horn neurons in which an oxytocin-receptor agonist enhanced glutamatergic and GABAergic but not glycinergic spontaneous transmissions. In order to know a developmental change and also sexual difference in the actions of oxytocin, we examined its effect on spontaneous synaptic transmission in adult female and young male rat SG neurons by using the whole-cell patch-clamp technique in spinal cord slices. In adult female rats, oxytocin produced an inward current at ?70 mV without a change in excitatory transmission. GABAergic and glycinergic transmissions were enhanced by oxytocin, the duration of which enhancement was much shorter than in adult male rats. In young (11–21 postnatal days) male rats, oxytocin produced not only an inward but also outward current at ?70 mV, and presynaptically inhibited or facilitated excitatory transmission, depending on the neurons tested; both GABAergic and glycinergic transmissions were enhanced by oxytocin. The inhibitory transmission enhancements in adult female and young male rats were sensitive to tetrodotoxin. Although the data may not be enough to be estimated, it is suggested that synaptic modulation by oxytocin in SG neurons, i.e., cellular mechanism for its antinociceptive action, exhibits a developmental change and sexual difference.     

Development of GABAergic connections in vitro: increasing efficacy of synaptic transmission is not accompanied by changes in miniature currents.

Development of inhibitory synaptic transmission was studied using a dissociated cell culture from the superior colliculus of neonatal rat. Patch-clamp recordings in the whole-cell configuration were performed to measure evoked (single-cell-activated) inhibitory postsynaptic currents (IPSCs), miniature IPSCs and current responses to maximal concentrations of exogenous gamma-aminobutyric acid (GABA). Over a period of 3 weeks in vitro (DIV3-24), the fraction of synaptically coupled neurons raised from 0% to 76%. Evoked IPSCs were first observed at DIV5. They had an average amplitude of 33.9 pA during the first week (n = 13) and 129.7 pA during the fourth week (n = 48). This increase by a factor of 3.8 represents a significant rise in the efficacy of GABAergic transmission during in vitro development. However, no developmental change has been observed in the average amplitudes of miniature somatic IPSCs. The latter remained at an average level of about 9 pA (symmetrical chloride concentration and a driving force of 68 mV). No increase was found also in whole-cell current densities induced by saturating concentrations of exogenous GABA. Our results suggest that under the given conditions, synapse maturation was primarily the result of presynaptic sprouting. This conclusion is further supported by bouton counts in immunostained collicular cultures, where the number of axosomatic and axodendritic GABAergic contacts per neuron increased from 0.54 and 0.37, respectively, at DIV3, to 13.84 and greater than 23.1, at DIV24. The overall density of GABAergic neurons decreased during this period from about 41,000/cm2 to 15,600 cm2, indicating that a growing number of contacts is formed by a declining number of presynaptic neurons.     

Opioid inhibition of GABA release from presynaptic terminals of rat hippocampal interneurons.

Opiates and the opioid peptide enkephalin can cause indirect excitation of principal cortical neurons by reducing inhibitory synaptic transmission mediated by GABAergic interneurons. The mechanism by which opioids mediate these effects on interneurons is unknown, but enkephalin hyperpolarizes the somatic membrane potential of a variety of neurons in the brain, including hippocampal interneurons. We now report a new, more direct mechanism for the opioid-mediated reduction in synaptic inhibition. The enkephalin analog D-Ala2-Met5-enkephalinamide (DALA) decreases the frequency of miniature, action potential-independent, spontaneous GABAergic inhibitory postsynaptic currents (IPSCs) without causing a change in their amplitude. Thus, we conclude that DALA inhibits the action potential-independent release of GABA through a direct action on interneuronal synaptic terminals. In contrast, DALA reduces the amplitude of action potential-evoked, GABA-mediated IPSCs, as well as decreases their frequency. This suggests that the opioid-mediated inhibition of non-action potential-dependent GABA release reveals a mechanism that contributes to reducing action potential-evoked GABA release, thereby decreasing synaptic inhibition.     

A long CAG repeat in the mouse Sca1 locus replicates SCA1 features and reveals the impact of protein solubility on selective neurodegeneration

A single nicotine exposure increases dopamine levels in the mesolimbic reward system for hours, but nicotine concentrations experienced by smokers desensitize nAChRs on dopamine neurons in seconds to minutes. Here, we show that persistent modulation of both GABAergic and glutamatergic synaptic transmission by nicotine can contribute to the sustained increase in dopamine neuron excitability. Nicotine enhances GABAergic transmission transiently, which is followed by a persistent depression of these inhibitory inputs due to nAChR desensitization. Simultaneously, nicotine enhances glutamatergic transmission through nAChRs that desensitize less than those on GABA neurons. The net effect is a shift toward excitation of the dopamine reward system. These results suggest that spatial and temporal differences in nicotinic receptor activity on both excitatory and inhibitory neurons in reward areas coordinate to reinforce nicotine self-administration.     

Expression of GABA transporters,GAT-1 and GAT-3, in the cerebral cortex and thalamus of the rat during postnatal development

The cellular and subcellular localization of two GABA transporters, GAT-1 and GAT-3, was investigated using immunocytochemical methods in the rat cerebral cortex and thalamus during postnatal development. The distribution of the transporters is compared with that of the neuronal marker GABA, and with that of vimentin and of glial fibrillary acidic protein, which identify immature and mature astrocytes, respectively. Our observations show that the two transporters are already expressed at birth in both brain areas with the same cellular localization as in adult rats, as GAT-1 is present in growth cones and terminals only in the cortex, whereas both transporters are expressed in astrocytes in the cortex and thalamus. The distribution of GAT-1 and GAT-3 undergoes postnatal changes reflecting in general the neurogenetic events of the neocortex and thalamus and, more specifically, the maturation of GABAergic innervation. The adult-like pattern of expression is achieved in the third postnatal week in the cortex and in the second postnatal week in the thalamus. The early expression of GAT-1 in GABAergic terminals confirms previous studies showing the existence of neuronal mechanisms of GABA uptake from the embryonic stages. As for the glial localization, the precocious existence of two astrocytic GABA transporters suggests that they operate through different functional mechanisms from birth, whereas their exclusively glial expression in the thalamus indicates that the astroglia plays a major role in the transport, recycling and metabolism of thalamic GABA.     

The Depolarizing Action of GABA Controls Early Network Activity in the Developing Hippocampus

Early in postnatal life γ-aminobutyric acid (GABA), the primary inhibitory transmitter in adults, excites targeted neurons by an outwardly directed flux of chloride which results from the unbalance between the cation–chloride cotransporters NKCC1 and KCC2, involved in chloride uptake and extrusion, respectively. This effect contributes to generate synchronized network activity or giant depolarizing potentials (GDPs) in the developing hippocampus. Here, we review some recent data concerning the mechanisms by which GDPs are generated and their functional role in enhancing synaptic efficacy at poorly developed GABAergic and glutamatergic synapses. In adulthood, reshaping neuronal circuits due to changes in chloride homeostasis and to the shift of GABA from hyperpolarizing to depolarizing, has been implicated in several neurological disorders, including epilepsy. Evidence has been recently provided that in chronically nerve growth factor-deprived mice expressing a progressive age-dependent neurodegenerative pathology resembling that observed in patients with Alzheimer’s disease, the reduced expression of mRNA encoding for the Kcc2 gene and the depolarizing action of GABA lead to the reorganization of the neuronal hippocampal network. This may represent a novel mechanism by which GABAergic signaling counterbalances the loss of synaptic activity in neurodegenerative diseases.     

Polysynaptic subcircuits in the neocortex: spatial and temporal diversity

Inhibitory pathways in the neocortex display a variety of temporal and spatial patterns, maintaining a dynamic balance with excitatory synaptic activity. Recent studies have revealed prevalent polysynaptic subcircuits within the neocortical microcircuitry. These subcircuits involve excitatory and inhibitory connections that are activated by neurons both in supragranular and infragranular cortical layers and mediated by different mechanisms. Interestingly, in these subcircuits inhibition is induced by discharge of pyramidal cells, and excitation is caused by specific types of GABAergic interneurons. The different polysynaptic subcircuits are discussed with respect to their spatial and temporal properties and their possible functional role in cortical processing.     

Anti-allodynic and promotive effect on inhibitory synaptic transmission of riluzole in rat spinal dorsal horn

Riluzole (2-amino-6-(trifluoromethoxy)benzothiazole) is a drug known for its inhibitory effect on glutamatergic transmission and its anti-nociceptive and anti-allodynic effects in neuropathic pain rat models. Riluzole also has an enhancing effect on GABAergic synaptic transmission. However, the effect on the spinal dorsal horn, which plays an important role in modulating nociceptive transmission, remains unknown. We investigated the ameliorating effect of riluzole on mechanical allodynia using the von Frey test in a rat model of neuropathic pain and analyzed the synaptic action of riluzole on inhibitory synaptic transmission in substantia gelatinosa (SG) neurons using whole-cell patch clamp recordings. We found that single-dose intraperitoneal riluzole (4 mg/kg) administration effectively attenuated mechanical allodynia in the short term in a rat model of neuropathic pain. Moreover, 300 μM riluzole induced an outward current in rat SG neurons. The outward current induced by riluzole was not suppressed in the presence of tetrodotoxin. Furthermore, we found that the outward current was suppressed by simultaneous bicuculline and strychnine application, but not by strychnine alone. Altogether, these results suggest that riluzole enhances inhibitory synaptic transmission monosynaptically by potentiating GABAergic synaptic transmission in the rat spinal dorsal horn.     

Class-specific features of neuronal wiring

Brain function relies on specificity of synaptic connectivity patterns among different classes of neurons. Yet, the substrates of specificity in complex neuropil remain largely unknown. We search for imprints of specificity in the layout of axonal and dendritic arbors from the rat neocortex. An analysis of 3D reconstructions of pairs consisting of pyramidal cells (PCs) and GABAergic interneurons (GIs) revealed that the layout of GI axons is specific. This specificity is manifested in a relatively high tortuosity, small branch length of these axons, and correlations of their trajectories with the positions of postsynaptic neuron dendrites. Axons of PCs show no such specificity, usually taking a relatively straight course through neuropil. However, wiring patterns among PCs hold a large potential for circuit remodeling and specificity through growth and retraction of dendritic spines. Our results define distinct class-specific rules in establishing synaptic connectivity, which could be crucial in formulating a canonical cortical circuit.