Mapping of spinal interneurons involved in regulation of the lower urinary tract in juvenile male rats.

Coordination between the urinary bladder (BL) and external urethral sphincter (EUS) is necessary for storage and elimination of urine. In rats interneuronal circuits at two levels of the spinal cord (i.e., L6-S1 and L3-L4) play an important role in this coordination. In the present experiments retrograde trans-synaptic transport of pseudorabies virus (PRV) encoding fluorescent markers (GFP and RFP) was used to trace these circuits. To examine the relative localization of EUS-related and BL-related interneuronal populations we injected PRV-GFP into the EUS and PRV-RFP into the BL wall. The PRV infected populations of spinal interneurons were localized primarily in the dorsal commissure (DCM) of L6/S1 and in a hypothesized lumbar spinal coordinating center (LSCC) in L3/L4 above and lateral to central canal (CC). At both sites colocalization of markers occurred in a substantial number of labeled interneurons indicating concomitant involvement of these double-labelled neurons in the EUS- and BL-circuits and suggesting their role in EUS-BL coordination. Intense GFP or RFP fluorescent was detected in a subpopulation of cells at both sites suggesting that they were infected earlier and therefore likely to represent first order, primary interneurons that directly synapse with output neurons. Larger numbers of weakly fluorescent neurons that likely represent second order interneurons were also identified. Within the population of EUS-related first order interneurons only 3-8 % exhibited positive immunoreaction for an early transcription factor Pax2 specific to GABAergic and glycinergic inhibitory neurons suggesting that the majority of interneurons in DCM and LSCC projecting directly to the EUS motoneurons are excitatory.

Spontaneous field potentials in the glomeruli of the olfactory bulb: the leading role of juxtaglomerular cells.

Field potentials recorded in the olfactory bulb glomerular layer (GL) are thought to result mainly from activation of mitral and tufted cells. The contribution of juxtaglomerular cells (JG) is unknown. We tested the hypothesis that JG are the main driving force to novel spontaneous glomerular layer field potentials (sGLFPs), which were recorded in rat olfactory bulb slices maintained in an interface chamber. We found that sGLFPs have comparable magnitudes, durations and frequencies both in standard horizontal slices, where all layers with all cell types were present, and in isolated GL slices, where only JG cells were preserved. Hence, the impact of mitral and deep/medium tufted cells to sGLFPs turned out to be minor. Therefore, we propose that the main generators of sGLFPs are JG neurons. We further explored the mechanism of generation of sGLFPs using a neuronal ensemble model comprising all types of cells associated with a single glomerulus. Random orientation and homogenous distribution of dendrites in the glomerular neuropil along with surrounding shell of cell bodies of JG neurons resulted in substantial spatial restriction of the generated field potential. The model predicts that less than 20% of sGLFP can spread from one glomerulus to an adjacent one. The contribution of JG cells to the total field in the center of the glomerulus is estimated as approximately 50% ( approximately 34% periglomerular and approximately 16% external tufted cells), whereas deep/medium tufted cells provide approximately 39% and mitral cells only approximately 10%. Occasionally, some sGLFPs recorded in adjacent or remote glomeruli were cross-correlated, suggesting involvement of interglomerular communication in information coding. These results demonstrate a leading role of JG cells in activation of the main olfactory bulb (MOB) functional modules. Finally, we hypothesize that the GL is not a set of independent modules, but it represents a subsystem in the MOB network, which can perform initial processing of odors.

Distribution of DNA, proteins, and lipids in cells of the olfactory bulb in rats of different ages.

Luminescence and absorption stains specific for DNA (acridine orange, ethidium bromide), proteins (silver nitrate), and lipids (Sudan III) were used to study the distribution of DNA, proteins, and lipids in sections of the olfactory bulb in rats, studies being performed after fixation of brains with paraformaldehyde. DNA was found to be more abundant in the glomerular cell layer than the mitral cell layer. Higher quantities of DNA were present in the granular layer, located beneath the mitral layer. The characteristics of cell layers in the olfactory bulb were studied in rats aged two days and one month. There were differences between the layers of rats of different ages in terms of the content and distribution of DNA, though there were no differences in the total protein or lipid contents. Glomeruli were immature in two-day-old rats.

[DNA, proteins and lipids distribution in the cells of olfactory bulbs of rats of various ages].

Using luminescent and absorption stains, specific for DNA (acridine orange, ethidium bromide), proteins (silver nitrate) and lipids (sudan III), the distribution of these substances was studied in the sections of rat olfactory bulbs, fixed by paraformaldehyde. DNA prevalence was found in glomerular cell layer as compared with the mitral one. Large amount of DNA was detected in granular cell layer, underlying the mitral one. The peculiarities of cellular layers of olfactory bulbs of 2-day-old rats were compared with those ones in 1-month-old animals. In rats of different ages, the differences were found in DNA content and distribution between layers, while no differences were detected in total protein and lipids. In 2-day-old rats glomerular underdevelopment was demonstrated.

[Neurogenesis in the mature olfactory bulb and it's possible functional destination]. / Neirogenez v zreloi oboniatel'noi lukovitse i ego vozmozhnoe funktsional'noe prednaznachenie.

This review is concerned with neurogenesis in the mature mammalian brain with emphasis on cell population renewal in the olfactory bulb (OB). The structural and functional features of the OB are considered along with data on neurotropic viruses and toxic dust penetration into the CNS through the OB. We hypothesize a protective role of neurogenesis in the mature OB. This suggests that normal renewal of cell populations in the OB is an important barrier mechanism protecting the brain from invasion of small amounts of harmful neurotropic agents (ex. viruses and particles of toxic dust), which can cause various neurodegenerative diseases.

Membrane and network theta-rhythm generation in hippocampal slices.

The hippocampal rhythms observed in vivo are the result of a complex interplay between cellular and synaptic properties within the hippocampus, and extra-hippocampal tonic as well as periodic inputs. For the stable rhythm to occur, the hippocampal circuitry should have the potential to oscillate at the specific frequencies. The in vitro studies revealed multiple mechanisms supporting the generation of the theta rhythm, which is the main operational mode of the hippocampus. In the hippocampus and related structures cellular membranes can oscillate at theta rhythm when they are depolarized to near-threshold membrane potentials; membranes are also adjusted to resonate with the external signal applied at theta frequency. Synaptically connected hippocampal network alone can generate theta rhythm when a necessary tonic excitation is provided. Finally, rhythmic inputs in theta range from the septum and entorhinal cortex have a propensity to synchronize oscillations in the whole hippocampal formation and associated structures to operate in a unified mode of activity. Based on the results obtained in slices and slice cultures, the present review shows this multilevel hierarchy, which serves to guarantee easy occurrence and reliable maintenance of the theta rhythm in the hippocampus.

Centre-surround inhibition among olfactory bulb glomeruli.

Centre-surround inhibition--the suppression of activity of neighbouring cells by a central group of neurons--is a fundamental mechanism that increases contrast in patterned sensory processing. The initial stage of neural processing in olfaction occurs in olfactory bulb glomeruli, but evidence for functional interactions between glomeruli is fragmentary. Here we show that the so-called 'short axon' cells, contrary to their name, send interglomerular axons over long distances to form excitatory synapses with inhibitory periglomerular neurons up to 20-30 glomeruli away. Interglomerular excitation of these periglomerular cells potently inhibits mitral cells and forms an on-centre, off-surround circuit. This interglomerular centre-surround inhibitory network, along with the well-established mitral-granule-mitral inhibitory circuit, forms a serial, two-stage inhibitory circuit that could enhance spatiotemporal responses to odours.

Cholinergic modulation of neuron spike responses to dendritic and somatic application of excitatory amino acids.

The effects of acetylcholine on the spike discharges of neurons induced by iontophoretic application of excitatory amino acids to the bodies and dendrites of cells were studied in 98 neurons in living slices of guinea pig parietal cortex. Acetylcholine applied microiontophoretically to both the bodies and dendrites facilitated improvements in the parameters of responses induced by dendritic activation, with significant decreases in latent periods and increases in the intensity and duration of responses. Thee effects were stably induced at distances of 300 microm from the body and lasted 1 min after exposure to acetylcholine ended. Responses induced by application of excitatory amino acids directly to the cell body did not change significantly in the presence of acetylcholine regardless of the point on the membrane at which they were applied. It is concluded that the predominant effect of acetylcholine is on the efficiency of dendrosomatic conduction.

[Cholinergic modulation of neuronal spike reactions to dendritic and somatic application of excitatory acids]. / Kholinergicheskaia moduliatsiia impul'snykh reaktsii neironov na dendritnoe i somaticheskoe podvedenie vozbuzhdaiushchikh aminokislot.

Acetylcholine effects on neuronal firing responses evoked by somatic or dendritic applications of excitatory amino acids were studied in slices of guinea-pig parietal cortex. Excitatory reactions initiated by dendritic activation were enhanced by acetylcholine wherever it was iontophoretically applied: either to soma or dendrites. The effect consisted in shortening spike response latencies and increasing response intensity and duration. The modified responses were recorded within 1-min interval after acetylcholine microinjections at a distance within 300 microns of the soma. Parameters of responses to somatic applications of excitatory amino acids were not significantly changed by acetylcholine. The results suggest that acetylcholine improves dendritic propagation rather than membrane excitability.

Cholinergic excitation of dendrites in neocortical neurons.

Discharge patterns were studied in response to iontophoretic application of acetylcholine to the soma and dendrites of 128 neocortical pyramidal neurons of layer V. Extracellular recordings were obtained from slices of the guinea-pig parietal cortex. All responses found were excitatory and were better expressed in spontaneously firing cells than in silent ones. Sensitivity to acetylcholine was approximately the same at somatic and dendritic sites in all the cells. Activation of muscarinic receptors gave rise to firing patterns with equal latencies and intensities when applied to both soma and dendrites. The latter suggests that membrane excitation elicited in dendrites by binding of acetylcholine to muscarinic cholinoreceptors is likely to propagate towards the soma through intracellular biochemical processes. Modulating effect of acetylcholine on output firing patterns, elicited by dendritic application of excitatory amino acids, included shortening of the somatic response latency and increase of response intensity and duration. We propose that, in contrast to glutamatergic excitation, the spread of cholinergic excitation along dendrites involves intra-cellular chemical signalling and results in changing the electrical properties of dendrites all over their length.

Responses of cortical neurons to microiontophoretic application of acetylcholine to their dendrites.

Spike responses of neurons to the microiontophoretic application of acetylcholine to the soma and the dendrites were studied. The somatic and dendritic membranes had virtually equal sensitivity to acetylcholine. Only activatory responses were seen, which were most typical of spontaneously active neurons. Muscarinic activation induced spike responses with equal latent periods and equal intensities on application of acetylcholine to dendrites and the soma. It is suggested that intracellular chemical signaling is involved in the propagation of cholinergic excitation via dendrites.

[The reactions of the cortical neurons to the microiontophoretic delivery of acetylcholine to their dendrites]. / Reaktsii neironov kory na mikroionoforeticheskoe podvedenie atsetilkholina k ikh dendritam.

In slices of parietal cortex of a guinea pig spike reactions were studied induced in neurons by iontophoretical applications of acetylcholine to their somata and dendrites. The results were obtained from 128 units. When applied to different sites of the neuronal membrane acetylcholine produced an increase in firing activity nearly in the same percent of cases (50-75%). The reactions to acetylcholine were most typical for spontaneously active neurons. The slow onset (to 8 sec) and long duration (to 25 sec) of responses evoked by acetylcholine point to an involvement of muscarinic receptors. Spike responses evoked by acetylcholine application to soma and dendrites were of the same latencies and magnitude. The most distant dendritic site where the acetylcholine excitation was able to evoke response of the soma was separated from it by 300-400 mcm. It is suggested that acetylcholine excitation propagates from dendritic points to the soma with intracellular biochemical processes.

Functional geometry of amino acid sensitive membrane of layer V neurons in the guinea-pig neocortex in vitro.

On guinea-pig neocortical slices the spatial organization of dendrites sensitive to excitatory amino acids was studied. Extracellular recording were obtained from the the soma of layer V neurons. Responses of 135 neurons to iontophoretically applied glutamate or aspartate have been analysed. An increased firing rate to somatic and most of dendritic applications were of short latency not exceeding 500 ms. Dendritic applications caused somatic responses with far longer latencies (up to 2-3 s) in 18% of cases. Latencies of responses to excitatory amino acids applied to several dendritic sites of the same neuron had similar values. The greatest reactions were obtained in response to excitatory amino acids imposed to the soma and proximal dendrites. At a distance of 100 microm beyond the soma in the basal region and region and further than 300 microm in the apical region excitatory amino acid applications produced two to three times less intensive somatic response. The area where dendritic activation gave rise to change in neuronal firing was confined to 350 and 800 microm for basal and apical dendrites, respectively. Topography of effective dendritic sites fell into the area corresponding to anatomically known outline of dendritic tree of pyramidal neurons. This fact implies that in our experiments we basically dealt with layer V pyramids. The results obtained suggest that local activation of distal dendrites may elicit spike generation in the soma. Different electrical properties of somatic and dendritic membranes are discussed.

Physiological properties of anatomically identified axo-axonic cells in the rat hippocampus.

1. The properties of a well-defined type of GABAergic local circuit neuron, the axo-axonic cell (n = 17), were investigated in rat hippocampal slice preparations. During intracellular recording we injected axo-axonic cells with biocytin and subsequently identified them with correlated light and electron microscopy. Employing an immunogold-silver intensification technique we showed that one of the physiologically characterized cells was immunoreactive for gamma-aminobutyric acid (GABA). 2. Axo-axonic cells were encountered in the dentate gyrus (n = 5) as well as subfields CA3 (n = 2) and CA1 (n = 10). They generally had smooth, beaded dendrites that extended throughout all hippocampal layers. Their axons ramified densely in the cell body layers and in the subjacent stratum oriens or hilus, respectively. Tested with electron microscopy, labeled terminals (n = 53) established synapses exclusively with the axon initial segment of principal cells in strata oriens and pyramidale and rarely in lower radiatum. Within a 400-microns slice a single CA1 axo-axonic cell was estimated to be in synaptic contact with 686 pyramidal cells. 3. Axo-axonic cells (n = 14) had a mean resting membrane potential of -65.1 mV, an average input resistance of 73.9 M omega, and a mean time constant of 7.7 ms. Action potentials were of short duration (389-microseconds width at half-amplitude) and had a mean amplitude of 64.1 mV. 4. Nine of 10 tested cells showed a varying degree of spike frequency adaptation in response to depolarizing current injection. Current-evoked action potentials were usually curtailed by a deep (10.2 mV) short-latency afterhyperpolarization (AHP) with a mean duration of 28.1 ms. 5. Cells with strong spike frequency accommodation (n = 5) had a characteristic firing pattern with numerous spike doublets. These appeared to be triggered by an underlying depolarizing afterpotential. In the same cells, prolonged bursts of action potentials were followed by a prominent long-duration AHP with a mean time constant of 1.15 s. 6. Axo-axonic cells responded to the stimulation of afferent pathways with short-latency excitatory postsynaptic potentials (EPSPs) or at higher stimulation intensity with up to three action potentials. Axo-axonic cells in the dentate gyrus could be activated by stimulating the CA3 area as well as the perforant path, whereas in the CA1 area responses were elicited after shocks to the perforant path, Schaffer collaterals, and the stratum oriens-alveus border. 7. In the CA1 area the EPSP amplitude increased in response to membrane hyperpolarization.(ABSTRACT TRUNCATED AT 400 WORDS)

Background firing activity in guinea-pig neocortex in vitro.

The background firing activity was recorded extracellularly in experiments on guinea-pig neocortical slices maintained in vitro. The following types of background firing activity were revealed: (i) high regular single spikes (48%), (ii) irregular single spikes (15%), (iii) bursts (7%), (iv) groups (7%), (v) mixed activity where single spikes alternated with bursts or groups (28%). The specific interspike interval distribution and the specific shape of autocorrelogram corresponded to each of these background firing activity types. Furie analysis of autocorrelograms showed periodic components in spike sequences with the maxima at 3, 12, and 28 Hz. When blocking synaptic transmission with 100 mM adenosine, about 70% of the background active cells "fell silent" and the remaining 30% of neurons continued to generate action potentials. The latter seem to be actual spontaneously active neurons, i.e. they were capable of autonomous spike generation. We failed to find any correlation between the type of neuronal firing and the ability of neurons to be spontaneously active. The selective blockade of inhibitory synapses with 100 mM picrotoxine did not practically change the character of background firing activity though the responses to stimulation became epileptic. An important conclusion to emerge from this study is that the background firing activity in cortical slices can include the actual spontaneous discharges related to intrinsic cell properties as well as those concerned with synaptic actions. Furthermore, a small number of spontaneously active neurons seem to be able to synaptically activate twice the number of cells. The inhibitory interneurons did not significantly influence the propagation of excitation with the absence of stimulation.

[The dependence of the impulse reactions in surviving cortex slices on the stimulation parameters]. / Zavisimost' impul'snykh reaktsii v perezhivaiushchikh srezakh kory ot parametrov stimuliatsii.

The dependence of evoked neuronal discharges on stimulus electrode position, on power, frequency and duration of stimulation was investigated in guinea-pig neocortical slices. At suprathreshold stimulus intensity and under low frequency (about 0.1/s) and limited duration of stimulus series (10-30) the discharge pattern was usually well preserved. A, more intensive, higher-frequency or too long stimulation often led to transient habituation of responses. When the distances from neuron to the stimulus sites were equal the radial propagation of excitation seemed to be easier than tangential one.

[Cytoplasmic RNA in neocortical neurons in vitro]. / Sostoianie tsitoplazmaticheskoi RNK v neironakh neokorteksa in vitro.

The investigation of the ratio of neuronal cytoplasmic RNA mono- and two-chain sections was performed in thin neocortical slices by means of fluorescent probing. Changes of the relative intensity of Acridine Orange red luminescence indicated a decrease in the quota of monochain sections in the common neuronal RNA pool. In eight hours of superfusion it was authentically less than in the control slices and in the slices which were incubated during four hours. Deep profiles of red fluorescence became plane in the process of incubation. It was suggested that the essential decrease in the red luminescence intensity of cytoplasmic RNA in vitro, observed between four and eight hours, may be conditioned by disintegration of polysomes to monosomes, and this, in its turn, may cause the decrease in ribosomal synthetic activity.

[The spread of excitation in surviving neocortical slices]. / Rasprostranenie vozbuzhdeniia v perezhivaiushchikh srezakh neokorteksa.

The propagation of excitation has been studied in neocortical guinea pig slices maintained in vitro. Stimuli were given at different distances from the chosen neuron. Comparison of latencies of the evoked impulse reactions indicated a complex structure and configuration of the vertically oriented ensemble of comparatively synchronously activated neurons. It is assumed that such an ensemble has a narrow (less than 300 microns) top layer II, a wide (approximately 600 microns and more) middle part in layer V and a narrowed (approximately 300 microns) basis in layer VI. A heterogeneity of the ensemble (the module) with respect to vertical propagation of excitation is found.

[Background impulsation of cortical neurons during blockade of synaptic inhibition in vital slices]. / Fonovaia impul'satsiia korkovykh neironov pri blokade sinapticheskogo tormozheniia v perezhivaiushchikh srezakh.

Investigation of the picrotoxin effect on the background neuronal activity was performed in guinea-pig cerebral cortex slices maintained in vitro. Extracellular recording showed that the blockade of GABA-ergic inhibition did not change characteristics of spike trains significantly, although the stimulus-evoked reactions became epileptic. These data give evidence that the inhibition processes in the background activity of cortex slices are expressed extremely slightly and have no intrinsic influence on this activity.

Prolonged excitation of individual neocortical neurons, structure of background impulse activity.

The background impulse activity of individual neurons was recorded extracellularly in the cerebral cortex of the cat during the prolonged microionophoretic delivery to these neurons of L-glutamate. Glutamate ionophoresis ensured the transition of the neuron to an elevated, but stable level of activation. An autocorrelation analysis of the trains of impulses showed that in spite of the multiple rise in the average discharge frequency, the type of background impulse activity and the periods of increased and reduced probability of discharges for the most part remained constant. The obtained data indicate that an individual cell is unable to influence substantially the interneurons connected with it, while the type of background activity of a neuron is determined primarily by the level of activation of the cellular ensemble incorporating this neuron.