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[Post-tetanic modification of the efficacy of excitatory transmission in neuronal networks with interhemispheric connections].

The modifiable reciprocal transcallosal monosynaptic excitatory connections were for the first time detected in vivo experiments in rat motor cortex using multiunit recording and crosscorrelation analysis, It was shown that high-frequency microstimulation (MCS) of a small group of cortical cells of one hemisphere produces long-term changes in the efficacy of transcallosal excitatory connections, and also ipsilateral connections in both hemispheres. The posttetanic changes appear as long-term potentiation (LTP) and long-term depression (LTD). The bursting neurons were found to have more favorable conditions for the induction of LTP of most converging inputs (in contrast to cells with other discharge patterns). Both LTP and LTD could be simultaneously induced in synapses formed by axon collaterals of a callosal cell on several neurons. LTP and LTD could be simultaneously obtained at diverse synapses of the same cell. The number of spontaneously active callosal neurons as well as the number and efficacy of transcallosal connections increased after the MCS, whereas the number and efficacy of ipsilateral connections decreased. Basing on these data we assume that the ipsilateral inhibition is more effective than the transcallosal inhibition. MCS results in the modification of the pattern of initially existing connections between numerous neurons of an ensemble including cells of both hemispheres.

Action Potentials↗

[Morphological analysis of the formation of cluster organization of neurons, forming the corticocortical connections in the cat visual cortex in early postnatal ontogenesis].

The development of cluster organization of neurons forming direct interzonal associative projections from striate cortex (area 17) to extrastriate area 21a in cat was studied by a technique of horseradish peroxidase retrograde transport. Reconstruction of cortical region containing retrogradely labeled neurons was performed using continuous serial sections of the brain. The analysis of spatial organization was done and quantitative characteristics of distribution pattern of initial corticocortical neurons in striate cortex in kittens were obtained. Interzonal connections were detected since 14th postnatal day. Neuronal cluster organization started forming after day 21 while its pattern became similar to that in the adult animal both qualitatively and quantitatively by the end of the 2nd postnatal month, that is somewhat later than it was previously thought. The data obtained also demonstrate the age-dependent decrease in the convergence of the striate cortex projections to area 21a associated with the reduction of the area, occupied by initial neurons.

Animals↗

Evidence for a monosynaptic connection between slowly adapting pulmonary stretch receptor afferents and inspiratory beta neurones.

The synaptic connection between slowly adapting pulmonary stretch receptor afferents and inspiratory neurones within a region ventral to the tractus solitarius was determined using intracellular recording and spike triggered averaging techniques. When the vagus nerve was stimulated at intensities eliciting a Hering-Breuer reflex, the difference in mean latency between centrally recorded action potentials of slowly adapting pulmonary stretch receptor afferents and e.p.s.p.s of inspiratory beta neurones was 0.2 ms. This difference is indicative of a monosynaptic connection. Extracellular single unit spikes of slowly adapting pulmonary stretch receptors recorded from the nodose ganglion were used to trigger the averaging of synaptic noise recorded from inspiratory neurones. A prominent wave of synaptic depolarization was observed in all inspiratory beta neurones even when a small number of sweeps were averaged. This depolarization was absent from inspiratory alpha neurones. The shape indices of these depolarizations are consistent with a monosynaptic connection between slowly adapting pulmonary stretch receptor afferents and inspiratory beta neurones. In addition, the data raise the possibility that this connection is multiple and distributed.

Animals↗

Constrained synaptic connectivity in functional mammalian neuronal networks grown on patterned surfaces.

The use of ordered neuronal networks in vitro is a promising approach to study the development and the activity of small neuronal assemblies. However, in previous attempts, sufficient growth control and physiological maturation of neurons could not be achieved. Here we describe an original protocol in which polylysine patterns confine the adhesion of cellular bodies to prescribed spots and the neuritic growth to thin lines. Hippocampal neurons in these networks are maintained healthy in serum free medium up to 5 weeks in vitro. Electrophysiology and immunochemistry show that neurons exhibit mature excitatory and inhibitory synapses and calcium imaging reveals spontaneous activity of neurons in isolated networks. We demonstrate that neurons in these geometrical networks form functional synapses preferentially to their first neighbors. We have, therefore, established a simple and robust protocol to constrain both the location of neuronal cell bodies and their pattern of connectivity. Moreover, the long term maintenance of the geometry and the physiology of the networks raises the possibility of new applications for systematic screening of pharmacological agents and for electronic to neuron devices.

Action Potentials↗

GABAergic amacrine cells in rat retina: immunocytochemical identification and synaptic connectivity.

GABAergic neurons have been identified in light and electron microscopic preparations of rat retina by an immunocytochemical localization of the GABA-synthesizing enzyme, glutamic acid decarboxylase (GAD). GAD-positive neuronal somata are found only in the inner and middle parts of the inner nuclear layer, and GAD-positive neuronal terminals are observed exclusively within the inner plexiform layer (IPL) and the outermost part of the ganglion cell layer. Dense aggregations of GAD-positive terminals alternate with less dense zones to form a lamination of the IPL. GAD-positive terminals contain pleomorphic synaptic vesicles and are the presynaptic elements of conventional synapses onto bipolar and amacrine cell processes, as well as onto the somata and dendrites of ganglion cells. In addition, GAD-positive terminals are postsynaptic to unstained bipolar terminals and are components of synaptic dyads where they occasionally appear to form reciprocal synapses with the bipolar terminals, and serial with unstained amacrine processes. Probable synaptic contacts between adjacent GAD-positive terminals also have been observed. Most of the synaptic input to GAD-positive terminals comes from bipolar cells, while the small remaining input mainly comes from other GAD-positive terminals. The synaptic output to GAD-positive terminals is greatest to bipolar cells, followed in decreasing order by GAD-negative amacrine cells, ganglion cells, and other GAD-positive cells. The total synaptic output of GAD-positive cells appears to be more than twice as great as the total input of these cells. The location of GAD-positive somata, the distribution of GAD-positive terminals, and the synaptic relationships formed by these terminals all indicate that amacrine cells are the only GABAergic neurons in rat retina. Our observations also indicate that not all amacrines are GABAergic and suggest that GABAergic neurons may be limited to a narrow field subclass of amacrine cell. The findings concerning the synaptic connections of GABAergic amacrines suggest that such cells are the first link in several divergent pathways from bipolar to ganglion cells and that they probably serve more than one function since they feed synaptic activity forward directly upon ganglion cells as well as back upon bipolar cells.

Animals↗

Conditionally immortalized neural progenitor cells grafted to the striatum exhibit site-specific neuronal differentiation and establish connections with the host globus pallidus.

The cell line RN33B has been reported to differentiate into neurons in a site-specific manner when grafted to the cortex and hippocampus of adult rats. To investigate the fate of RN33B cells in a subcortical structure, we grafted RN33B cells into the intact or excitoxically lesioned striatum of adult or neonatal rats. The total number and phenotypic characteristics of the [3H]thymidine-labeled grafted cells were analyzed at different time points after transplantation. Transplanted RN33B cells were found to survive, integrate, and differentiate into both neurons and astrocytes, and a significant proportion of the cells (approx. 10%) were found to differentiate into cells with morphological and phenotypic characteristics of medium-sized striatal projection neurons. Retrograde tracing showed that at least some of the graft-derived neurons were capable of establishing connections with one of the primary striatal targets, the globus pallidus. These findings demonstrate a remarkable capacity of the RN33B cells for site-specific neuronal differentiation in both the adult and the developing striatum and suggest that the same differentiating factors that are operating during normal neurogenesis in brain development are retained, at least to some extent, also in the adult CNS.

Adult↗

The respiratory drive to thoracic motoneurones in the cat and its relation to the connections from expiratory bulbospinal neurones.

The descending control of respiratory-related motoneurones in the thoracic spinal cord remains the subject of some debate. In this study, direct connections from expiratory bulbospinal neurones to identified motoneurones were investigated using spike-triggered averaging and the strengths of connection revealed were related to the presence and size of central respiratory drive potentials in the same motoneurones. Intracellular recordings were made from motoneurones in segments T5-T9 of the spinal cord of anaesthetized cats. Spike-triggered averaging from expiratory bulbospinal neurones in the caudal medulla revealed monosynaptic EPSPs in all groups of motoneurones, with the strongest connections to expiratory motoneurones with axons in the internal intercostal nerve. In the latter, connection strength was similar irrespective of the target muscle (e.g. external abdominal oblique or internal intercostal) and the EPSP amplitude was positively correlated with the amplitude of the central respiratory drive potential of the motoneurone. For this group, EPSPs were found in 45/83 bulbospinal neurone/motoneurone pairs, with a mean amplitude of 40.5 microV. The overall strength of the connection supports previous measurements made by cross-correlation, but is about 10 times stronger than that reported in the only previous similar survey to use spike-triggered averaging. Calculations are presented to suggest that this input alone is sufficient to account for all the expiratory depolarization seen in the recorded motoneurones. However, extra sources of input, or amplification of this one, are likely to be necessary to produce a useful motoneurone output.

Action Potentials↗

Grafted septal neurons form cholinergic synaptic connections in the dentate gyrus of behaviorally impaired aged rats.

A group of aged, behaviorally impaired rats received suspension grafts of fetal septal-diagonal band tissue into the otherwise intact hippocampal formation. Three months after grafting, behaviorally recovered rats were studied by immunocytochemistry by using monoclonal antibodies to choline acetyltransferase (ChAT) and electron microscopy. The innervation of the host dentate gyrus by graft-derived ChAT-positive fibres was unmasked by acute removal of the intrinsic septal cholinergic innervation by fimbria-fornix transection 5-7 days before perfusion. The pattern of termination and ultrastructural connectivity were compared with previous results obtained from both young nonoperated control animals and a group of young rats with intrahippocampal septal grafts that had been subjected to denervation of the intrinsic cholinergic input at the time of transplantation. Graft-derived, ChAT-immunoreactive terminals formed abundant synaptic specializations with neuronal elements in the host dentate gyrus. The predominant postsynaptic target (about 65% of all boutons) was dendritic shafts, whereas about 20% of the boutons contacted dendritic spines. Very few synapses onto neuronal perikarya were found in the grafted aged rats. In some of these cases, however, it was possible to identify the target as dentate granule cells. This situation is very similar to that seen in young control rats but significantly different than the distribution observed in the denervated young grafted group, where axosomatic contacts predominated. The results indicate that the graft-induced behavioral improvement seen in the aged rats may depend on the formation of functional cholinergic graft connections with neuronal elements in the host hippocampal formation.

Aging↗

The contribution of GABA-ergic neurons to horizontal intrinsic connections in upper layers of the cat's striate cortex.

The contribution of neurons containing gamma-aminobutyric acid (GABA) to horizontal intrinsic projections in layers I-III of cat's striate cortex was investigated by combining GABA-immunohistochemistry with axonal tracing. After intracortical injections of Rhodamine-labelled latex microspheres Rhodamine-labelled neurons form patch- or bandlike aggregations (clusters) separated from each other by regions containing fewer, evenly distributed or no labelled neurons. Of the Rhodamine-labelled neurons about 5% display GABA-immunoreactive material (double labelled = DL-neurons). Approximately 70% of the DL-neurons occur at distances of less than 1 mm, and the remaining 30% at distances between 1 mm and 2.5 mm from the injection. About 60% of the DL-neurons reside within clusters and 40% are located in regions between clusters; the respective percentages of the Rhodamine labelled GABA-negative neurons are about 85 and 15. Considering their small number and their spatial distribution inhibitory interneurons seem to make only minor contributions to the clustered pattern of intrinsic connections. Our results demonstrate that the topographical organization of neurons giving origin to lateral inhibitory interactions in upper layers of cat's striate cortex is different from that of neurons mediating excitatory functions.

Animals↗

Central connections of the toad neural lobe as shown by retrograde neuronal labeling: classical and new connections.

Iontophoretic deposit of horseradish peroxidase (HRP) in the hypophysial neural lobe (NL) of the toad labeled neurons in the classical neurosecretory preoptic magnocellular nucleus. Further, a number of labeled neurons appeared in the whole extension of the periventricular and medial preoptic nuclei. The latter were continuous dorsally with two other labeled areas known as the bed nucleus of the hippocampal commissure (which appears to be homologous to the subfornical organ in mammals) and the ventromedial thalamic area of Frontera. The possible functional role of many of these afferents to the neural lobe is unknown because all types of neurons were labeled in the projecting zones and, in addition, many are outside of the generally recognized neurosecretory groups.

Afferent Pathways↗

The role of the brain in metamorphosis of the olfactory epithelium in the frog, Xenopus laevis.

Retrograde signaling from the brain to the olfactory sensory epithelium is important for neuronal survival, but the importance of the olfactory bulb in retrograde signaling during the naturally-induced, neuronal plasticity occurring during metamorphosis is unclear. The olfactory system of the African clawed frog (Xenopus laevis) undergoes dramatic rearrangements during metamorphosis, making this an ideal system in which to examine interactions between the brain and the olfactory sensory epithelium. The main olfactory epithelium of larvae, located in the principal cavity (PC), changes at metamorphosis in function, receptor neuron morphology, biochemistry, and axon termination sites. A new, "middle", cavity forms during metamorphosis that assumes all the characteristics of the larval PC. Using a combination of bulbectomy and olfactory transplantation, we investigated changes in expression of a marker protein (E7) and in apical ultrastructure in olfactory receptor neurons either (1) connected to the olfactory bulb, (2) connected to non-olfactory brain regions, or (3) with no apparent central nervous system (CNS) connections. We find that neurons in the middle cavity (MC) lacking connections with the CNS appear mature but neurons in the PC do not. Supporting cells in the PC undergo the changes normally observed during metamorphosis. Neurons connected to non-olfactory brain regions, either after bulbectomy or transplantation, appeared normal with regard to the changes normally expected after metamorphosis. These results suggest that influence from the brain is necessary for metamorphic changes in the X. laevis olfactory epithelium, but that these signals are not confined to the olfactory bulb; non-olfactory brain regions can also support these metamorphic changes.

Animals↗

Connections of hypothalamic paraventricular neurons with the dorsal medial thalamus and neurohypophysis: an electrophysiological study in the rat.

Connections between the hypothalamic paraventricular nucleus (PVN) and thalmic paraventricular nucleus ( PVT ) were examined using electrophysiological methods. Efferent projections of adjacent PVN cells were defined on the basis of antidromic activation from either PVT (n = 12) or neurohypophyseal (n = 38) stimulation; antidromic activation from both sites (n = 3) suggested that some PVN cells project both to the PVT and to the neurohypophysis. PVT stimulation evoked only weak orthodromic responses from 21% of PVN neurohypophyseal neurons, whereas short latency, high probability orthodromic responses were observed from 43% of PVN non-neurosecretory neurons. These data indicate reciprocal PVN- PVT connections and suggest that PVT afferents preferentially innervate non-neurosecretory PVN cells.

Animals↗

Transneuronal transfer of herpes simplex virus type 1 (HSV 1) from mixed limb nerves to the CNS. I. Sequence of transfer from sensory, motor, and sympathetic nerve fibres to the spinal cord.

The time course of transneuronal transfer of Herpes simplex virus type 1 (HSV 1) from sensory, motor, and sympathetic nerve fibres to connected spinal neurones was examined. After injection of a constant number of infectious units into distal forelimb or hindlimb nerves of inbred rats of the same age, the extent of viral transfer was strictly dependent on the survival time postinoculation (p.i.). Retrograde transport to somatic motoneurones occurred at 28-29 hours p.i. (stage 1), in synchrony with anterograde transneuronal transfer via small cutaneous afferents (to laminae I-II). At 36-43 hours p.i. (stage 2), retrograde transneuronal transfer from sympathetic nerve fibres first labelled sympathetic preganglionic neurones. At 48-51 hours p.i. (stage 3), transfer via sensory and sympathetic axons became more extensive, labelling laminae III-IV and other preganglionic neurones. Transneuronal transfer from large muscle afferents and motoneurones (to Clarke's columns and the spinal intermediate zone) occurred only at 66-78 hours p.i. (stage 4). Further increases in distribution (stages 5-6) obtained between 78 and 97 hours p.i. may reflect both specific labelling of second and third order neurones and a gradual local loss of specificity. These results indicate that transfer of HSV 1 occurs through all main classes of peripheral axons, but that both anterograde and retrograde transneuronal transfer from small (unmyelinated and fine myelinated) cutaneous and sympathetic axons precedes transfer from large (myelinated) cutaneous and muscle afferents and motor axons. Analysis of viral transfer at sequential intervals is required to distinguish serially connected neurones, determine the route of labelling, and ensure its specificity.

Animals↗

The molecular diversity of Dscam is functionally required for neuronal wiring specificity in Drosophila.

Alternative splicing of Dscam generates an enormous molecular diversity with maximally 38,016 different receptors. Whether this large diversity is required in vivo is currently unclear. We examined the role of Dscam in neuron-target recognition of single mechanosensory neurons, which connect with different target cells through multiple axonal branches. Analysis of Dscam null neurons demonstrated an essential role of Dscam for growth and directed extension of axon branches. Expression of randomly chosen single isoforms could not rescue connectivity but did restore basic axonal extension and rudimentary branching. Moreover, two Dscam alleles were generated that each reduced the maximally possible Dscam diversity to 22,176 isoforms. Reduction of Dscam diversity resulted in specific connectivity defects of mechanosensory neurons. Furthermore, the observed allele-specific phenotypes suggest functional differences among isoforms. Our findings provide evidence that a very large number of structurally unique receptor isoforms is required to ensure fidelity and precision of neuronal connectivity.

Alternative Splicing↗

Giant Aplysia neuron R2 reliably forms strong chemical connections in vitro.

The giant cholinergic neuron R2 of Aplysia was cultured in combination with identified neurons L11 and R15 and members of a group of left upper quadrant (LUQ) cells L2 to L6 from the abdominal ganglion. All of these neurons receive cholinergic input from other cells in vivo, but not from R2. In vitro, R2 reliably formed unidirectional chemical connections with these cells. Single action potentials in R2 produced a dual fast and slow inhibitory response in LUQ cells (L2 to L6), a dual fast inhibitory-slow excitatory response in L11, and a slow inhibitory response in R15. The connections formed on LUQ cells were characteristic of their cholinergic input, but the R2-L11 and the R2-R15 connections also had noncholinergic properties. Thus, unlike L10 which forms connections only with its normal targets in vitro, R2 forms strong chemical connections with other neurons which are not found in vivo. The properties of the R2 connections also suggest that it may release another neurotransmitter besides acetylcholine.

Action Potentials↗

Peripheral target specification of synaptic connectivity of muscle spindle sensory neurons with spinal motoneurons.

The source of environmental cues determining the central connections of muscle sensory neurons was investigated by manipulating chick embryos so that sensory neurons supplied a duplicate set of dorsal thigh muscles. These neurons projected out ventral nerve pathways and along motor axons that normally project to ventral muscles but their ultimate target tissue was the duplicate set of dorsal muscles. The central connections of these sensory neurons to motoneurons supplying normal dorsal muscles were then determined with intracellular recordings in isolated spinal cord preparations. Sensory neurons supplying individual duplicate dorsal muscles made the same connections as those supplying the corresponding normal dorsal muscles; the pattern of these connections was different than that made by afferents supplying ventral muscles. Sensory neurons thus made synaptic connections appropriate for their target muscle rather than for their more proximal ventral environment. These findings suggest that the target muscle is the source of cues that determine the central connections of the sensory neurons projecting to it. Motoneurons forced to innervate novel muscle received many of the same sensory inputs they would normally receive, suggesting that motoneurons are less influenced by their target tissue than sensory neurons.

Animals↗

Cross-correlation analysis of neuronal activities.

Nerve impulses are generally regarded as spike train and analyzed by use of various kinds of so-called time series analyses. Cross-correlation analysis is used to reveal temporal and/or spatial relationships between more than two spike trains in the neuronal circuit, in which two neurons are synaptically connected or two neurons are independent but receive a common input. Three main characteristics of discharge dependency between two neurons become clear from the cross-correlation histogram: 1) the direction of dependency; 2) the latency of interaction; and 3) the type of functional connections between the neurons. We reviewed the cross-correlation analysis from these points of view: 1) what is cross-correlation analysis; 2) what are primary and secondary effects; 3) what kinds of functional information can be obtained from the shape of the primary effect; 4) how sensitively cross-correlation can detect neuronal interaction; 5) how to express quantitatively the primary effect; 6) what kinds of extension technique are available from cross-correlation; and 7) research trends using cross-correlation analysis.

Animals↗