Interneuronal Signaling Research Articles

Vasopressin and oxytocin release within the brain: a dynamic concept of multiple and variable modes of neuropeptide communication

As exemplified particularly with vasopressin and oxytocin, release of neuropeptides within the brain occurs from dendrites, somata, and axons of neurosecretory neurons; mechanisms include activation of intracellular Ca 2+ stores, changed strength of synaptic input and altered interaction between transcription factors and gene promoters. Upon demand, both diffuse spread of neuropeptides in the extracellular fluid following dendritic release and focal release from axonal terminals may contribute to regionally and temporally varying combinations of neuromodulator and neurotransmitter actions, thus providing a theoretically unlimited variability in interneuronal signaling. Thus, instead of favoring volume or synaptic transmission following central neuropeptide release, a more dynamic concept is presented with multiple and variable modes of release and communication. This concept considers neuropeptides in the extracellular fluid of the brain rather than those in the cerebrospinal fluid or plasma as primary signals, triggering a variety of receptor-mediated effects, including those underlying behavioral and neuroendocrine regulation and psychopathology.

Activity-dependent homeostatic specification of transmitter expression in embryonic neurons

Neurotransmitters are essential for interneuronal signalling, and the specification of appropriate transmitters in differentiating neurons has been related to intrinsic neuronal identity and to extrinsic signalling proteins. Here we show that altering the distinct patterns of Ca2+ spike activity spontaneously generated by different classes of embryonic spinal neurons in vivo changes the transmitter that neurons express without affecting the expression of markers of cell identity. Regulation seems to be homeostatic: suppression of activity leads to an increased number of neurons expressing excitatory transmitters and a decreased number of neurons expressing inhibitory transmitters; the reverse occurs when activity is enhanced. The imposition of specific spike frequencies in vitro does not affect labels of cell identity but again specifies the expression of transmitters that are inappropriate for the markers they express, during an early critical period. The results identify a new role of patterned activity in development of the central nervous system.

Distance-Dependent Scaling of Calcium Transients Evoked by Backpropagating Spikes and Synaptic Activity in Dendrites of Hippocampal Interneurons

Although interactions between backpropagating action potentials and synaptic stimulations have been extensively studied in pyramidal neurons, dendritic propagation and the summation of these signals in interneurons are not nearly as well known. In this study, two-photon imaging was used to explore the basic properties of dendritic calcium signaling in CA1 stratum radiatum interneurons. In contrast to hippocampal pyramidal neurons, the backpropagating action potential-evoked calcium transients in dendrites of interneurons underwent a distance-dependent increment. Although, in proximal dendrites, an increment could be attributed to a smaller dendrite diameter, distal dendrites did not show such dependence. Calcium responses in interneurons had a smaller amplitude, slower rise time, and decay than in pyramidal neurons. To explore the factors underlying the difference, we compared the calcium-binding capacity in interneurons and in pyramidal neurons. Our finding that endogenous calcium buffers had a higher level in interneurons may primarily explain the different kinetics and amplitudes of calcium transients. Synaptic stimulation-evoked calcium transients were also larger at distant dendritic locations. The spread of these signals was restricted to 12-13 microm long dendritic compartments. Supporting the reported lack of long-term potentiation in these interneurons, we found only sublinear or linear summations of calcium responses to coincident synaptic inputs and backpropagating spikes.

Intracellular Ca 2+ microdomain-triggered exocytosis in neuroendocrine cells

Colocalization of voltage-gated Ca 2+ channels and exocytotic sites at the active zones of nerve terminals underlies ‘synchronous’ action potential discharge and synaptic vesicle exocytosis, thus allowing fast interneuronal signalling. Such a demand for a rapid release is not expected in neuroendocrine cells whose secretory products act throughout the entire organism. Nevertheless, by using evanescent field imaging of near-membrane Ca 2+ concentrations and fluorescently labelled vesicles, Becherer et al. have recently reported exocytosis of individual large dense-core vesicles triggered by Ca 2+ microdomains formed around clusters of open L-type Ca 2+ channels in chromaffin cells from the adrenal medulla. This finding, besides illustrating the power of new microscopy imaging techniques, directly demonstrates in neuroendocrine cells a functional interaction between Ca 2+ channels and secretory vesicles very much reminiscent of that in neurons.

Cellular accumulation of anandamide: consensus and controversy.

The endocannabinoids N-arachidonylethanolamine (AEA or anandamide) and 2-arachidonylglycerol (2-AG) are hypothesized to function in the brain as interneuronal signaling molecules. Prevailing models of the actions of these molecules require that they traverse cellular plasma membranes twice; first, following cellular synthesis and second, prior to enzymatic hydrolysis. The transmembrane movement of AEA has been studied in multiple laboratories with a primary focus on its cellular accumulation following extracellular administration. Although there are areas of consensus among laboratories regarding AEA accumulation, several aspects are very unclear. In particular, there is a lack of consensus in the literature regarding the importance of AEA hydrolysis by fatty acid amide hydrolase in maintaining the driving force for accumulation. Furthermore, evidence for and against a transmembrane carrier protein has been published. We have reviewed the available literature and present a working model of the processes that are involved in the cellular accumulation of AEA. It is our hypothesis that transmembrane movement of AEA is regulated by concentration gradient between extracellular and intracellular free AEA. Furthermore, it is our view that a significant portion of the intracellular AEA in most cells is sequestered either by a protein or lipid compartment and that AEA sequestered in this manner does not equilibrate directly with the extracellular pool. Finally, we discuss the available data that have been presented in support of a transmembrane carrier protein for AEA.

Glutamatergic modulation of GABAergic signaling among hippocampal interneurons: novel mechanisms regulating hippocampal excitability.

Because interneurons play a central role in regulating the excitability of the hippocampal formation, it is important to understand the mechanisms that modulate gamma-aminobutyric acid (GABA)ergic signaling among them. This study addresses the modulation of GABA release from interneuron terminals by presynaptic glutamate receptors. Whole-cell recordings were obtained from CA1 stratum radiatum interneurons in guinea pig hippocampal slices. Selective agonists and blockers of glutamate receptors were used to study modulation of GABAergic transmission by group III metabotropic receptors or kainate receptors. Antidromic action-potential initiation also was analyzed by stimulating the axons of interneurons. Agonists of group III metabotropic glutamate receptors attenuated monosynaptic GABAergic signals in interneurons, but not in pyramidal neurons, in agreement with anatomic evidence on the distribution of these receptors. Submicromolar kainate enhanced the frequency and amplitude of spontaneous GABAergic signals in interneurons. Kainate also depolarized the axons of hippocampal interneurons, and triggered spontaneous ectopic action potentials in axons. Synaptically released glutamate reproduced many of the effects of both agonists, implying that these receptors can sense the ambient glutamate concentration, and therefore indirectly respond to the excitatory traffic in the hippocampus. When the two classes of receptors were stimulated simultaneously, complex interactions were obtained. Group III metabotropic receptors and kainate receptors profoundly affect GABAergic signaling among interneurons of the hippocampus. Glutamatergic modulation of GABAergic signaling among interneurons represents a novel class of mechanisms that potentially plays a major role in determining the initiation, propagation, and termination of seizures.

Molecular pharmacology of the Na+-dependent transport of acidic amino acids in the mammalian central nervous system.

The Na+-dependent transport of L-glutamate (GluT) has been identified in brain tissue more than thirty years ago. Neurochemical studies, performed in various experimental models during 1970's, defined the basic rules for the selection or synthesis of GluT-specific substrates and inhibitors. The protein molecules (transporters) that mediate the translocation of the substrates across the plasma membrane have been cloned and studied during the last ten years. The sites on the transporters that bind the substrates favour glutamate-like or aspartate-like molecules with one positively charged and two negatively charged ionised groups. Substituents at C3 and C4 are often tolerated but substitutions at C2 or alterations of the ionisable groups usually impede the binding. The substrate binding sites display an "anomalous" selectivity towards stereoisomers. These structural requirements are shared by all Na+-dependent glutamate transporters thus making the design of transporter-selective ligands a challenging task. Moreover, the molecular mechanisms of the transport have not yet been adequately elucidated. Data from a wide variety of experimental studies strongly indicate that Na+-dependent GluT regulates the functioning of the glutamatergic excitatory synapses-the most important rapid inter-neuronal signalling system in the mammalian brain. Altered structural and/or functional properties of the Na+-dependent glutamate transporters have been implicated in the damage to the brain tissue following cerebral ischaemia and in the progressive loss of neurons in conditions such as Alzheimer dementia and amyotrophic lateral sclerosis. Furthermore, it seems that fine-tuning of glutamatergic neurotransmission by regulating the Na+-dependent GluT could be useful in the therapy of schizophrenia.

Interneuronal signalling is involved in induction of collateral sprouting of nociceptive axons

Collateral sprouting of cutaneous nociceptive axons into the adjacent denervated skin critically depends on the nerve growth factor, presumably originating from the degenerated neural pathways and denervated skin. We hypothesised that the degenerated neural pathways are necessary, but not sufficient, to induce collateral sprouting of nociceptive axons, and, in addition, that the interaction between the injured and non-injured neurones within a dorsal root ganglion can trigger sprouting of nociceptive axons also in the absence of the denervated skin. End-to-side nerve anastomosis, made in female Wistar rats by suturing the end of an excised peroneal nerve segment to the side of the intact sural nerve, was used as a model for sprouting which allowed us to study the putative induction mechanisms separately. If the nerves adjacent to the sural nerve were transected concomitantly with the coaptation of the end-to-side anastomosis, robust nociceptive axon sprouting into the anastomosed nerve segment was observed by the nerve pinch test and counting of myelinated axons. Collateral sprouting did not occur, however, either if the cells in the anastomosed nerve segment were killed by freezing and thawing, or if the adjacent nerves had not been injured. However, if the ipsilateral dorsal cutaneous nerves, having their neurones in the same dorsal root ganglia as the sural nerve, were transected, but no other nerves were injured, then the sural nerve axons sprouted in abundance through the anastomosis even in the absence of denervated skin around the sural nerve terminals. From these results we suggest that cells (probably proliferating Schwann cells) in the degenerated neural pathways are necessary but not sufficient to induce collateral sprouting of nociceptive axons, and that interactions between the injured and non-injured neurones within the dorsal root ganglion (i.e. direct or indirect interneuronal signalling) are important in this regard.

Chronic Blockade of Glutamate Receptors Enhances Presynaptic Release and Downregulates the Interaction between Synaptophysin-Synaptobrevin–Vesicle-Associated Membrane Protein 2

During development of neuronal circuits, presynaptic and postsynaptic functions are adjusted in concert, to optimize interneuronal signaling. We have investigated whether activation of glutamate receptors affects presynaptic function during synapse formation, when constitutive synaptic vesicle recycling is downregulated. Using primary cultures of hippocampal neurons as a model system, we have found that chronic exposure to both NMDA and non-NMDA glutamate receptor blockers during synaptogenesis produces an increase in miniature EPSC (mEPSC) frequency, with no significant changes in mEPSC amplitude or in the number of synapses. Enhanced synaptic vesicle recycling, selectively in glutamatergic nerve terminals, was confirmed by the increased uptake of antibodies directed against the lumenal domain of synaptotagmin. No increased uptake was detected in neuronal cultures grown in the chronic presence of TTX, speaking against an indirect effect caused by decreased electrical activity. Enhanced mEPSC frequency correlated with a reduction of synaptophysin-synaptobrevin-vesicle-associated membrane protein 2 (VAMP2) complexes detectable by immunoprecipitation. Intracellular perfusion with a peptide that inhibits the binding of synaptophysin to synaptobrevin-VAMP2 induced a remarkable increase of mEPSC frequency in control but not in glutamate receptor blocker-treated neurons. These findings suggest that activation of glutamate receptors plays a role in the downregulation of the basal rate of synaptic vesicle recycling that accompanies synapse formation. They also suggest that one of the mechanisms through which this downregulation is achieved is an increased interaction of synaptophysin with synaptobrevin-VAMP2.

Brain-derived neurotrophic factor modulates dendritic morphology of cerebellar basket and stellate cells: an in vitro study

The dendrites of cerebellar basket/stellate cells show a highly stereotyped orientation relative to granule cell axons (parallel fibers) and Purkinje cell dendrites. This specific morphology is acquired during the early postnatal phase of cerebellar development, when basket/stellate cells become synaptically integrated with Purkinje neurons and granule cells. In the present study, we used primary cerebellar cultures to test how the spatial arrangement of granule cell axons affects basket/stellate cell dendritic morphology. In addition, we sought to determine whether active signals as might be provided by granule cells, i.e. synaptic input and the neurotrophin, brain-derived neurotrophic factor, affect basket/stellate cell development. Our results confirm the critical role of parallel fiber orientation for basket/stellate dendritic morphogenesis. Moreover, we found that both electrical activity and brain-derived neurotrophic factor increased basket and stellate cell dendritic arborization. Together with previously published findings, our data led to the conclusion that both structural cues and active interneuronal signaling collaborate to bring about the precise morphogenesis of cerebellar basket/stellate cells. The distinct responses of various cerebellar phenotypes towards the morphogenetic effects of brain-derived neurotrophic factor suggest that this neurotrophin, within the developing cerebellum, enhances synaptic connectivity by concerting the formation of appropriate pre- and postsynaptic structures.

Horizontal cells reveal cone type-specific adaptation in primate retina.

The human cone visual system maintains contrast sensitivity over a wide range of ambient illumination, a property known as light adaptation. The first stage in light adaptation is believed to take place at the first neural step in vision, within the long, middle, and short wavelength sensitive cone photoreceptors. To determine the properties of adaptation in primate outer retina, we measured cone signals in second-order interneurons, the horizontal cells, of the macaque monkey. Horizontal cells provide a unique site for studying early adaptational mechanisms; they are but one synapse away from the photoreceptors, and each horizontal cell receives excitatory inputs from many cones. Light adaptation occurred over the entire range of light levels evaluated, a luminance range of 15-1,850 trolands. Adaptation was demonstrated to be independent in each cone type and to be spatially restricted. Thus, in primates, a major source of sensitivity regulation occurs before summation of cone signals in the horizontal cell.

Anesthetic-induced alteration of Ca2+ homeostasis in neural cells: a temperature-sensitive process that is enhanced by blockade of plasma membrane Ca2+-ATPase isoforms.

Many inhalation anesthetics at clinically relevant concentrations inhibit plasma membrane Ca2+-adenosine triphosphatase (PMCA) ion pumping in brain synaptic membranes and in cultured cells of neural origin. In this study, the authors investigated the effect of inhalation anesthetics on cytosolic calcium homeostasis in cortical neurons maintained at physiologic and room temperatures and on cortical neurons and pheochromocytoma cells with antisense blockade of specific PMCA isoforms. Using Ca2+-specific confocal microfluorimetry, the anesthetic effects on Ca2+ dynamics were examined in mouse embryonic cortical neurons in association with ligand-stimulated Ca2+ influx. Studies were done at 21 degrees C and 37 degrees C. Mouse embryonic cortical neurons with oligodeoxyribonucleotide blockade of PMCA2 expression and transfected rat pheochromocytoma cells with blocked expression of PMCA1 were also examined. Baseline and poststimulation peak cytosolic calcium concentrations ([Ca2+]i) were increased, and Ca2+ clearance was delayed in cells exposed at 37 degrees C, but not at 21 degrees C, to concentrations < or = 1 minimum alveolar concentration (MAC)-equivalent of halothane, isoflurane, and sevoflurane. Neurons exposed to xenon solutions < or = 0.4, 0.6, and 0.8 MAC showed dose-related perturbations of cytosolic Ca2+. Calcium dynamics were altered in neural cells with blocked PMCA isoform production, but at much lower halothane concentrations: 0.5 MAC for cortical neurons and 0.1 MAC for pheochromocytoma cells. By extruding Ca2+ through the plasma membrane, PMCA maintains resting neuronal [Ca2+]i at low levels and clears physiologic loads of Ca2+ after influx through calcium channels. Inhalation anesthetics perturb this process and thus may interfere with neurotransmitter release, altering interneuronal signaling.

Light enhances hydrodynamic signaling in the multimodal caudal photoreceptor interneurons of the crayfish

1. The caudal photoreceptor (CPR) interneurons in the sixth abdominal ganglion of the crayfish are complex, multi-modal interneurons. These cells respond directly to light with tonic spike discharges, and they integrate synaptic input from an array of fili-form mechanoreceptors on the tailfan. They also provide input to rostral command centers, inducing backward walking at high firing frequencies, and thus directly influence behavior. 2. We recorded CPR activity in response to weak hydrodynamic stimulation of the tailfan mechanoreceptors while under varying intensities of light shined on the sixth ganglion. Spike trains were characterized according to the mean discharge rate (MDR) and the power spectrum from which the signal-to-noise ratio (SNR) was calculated. 3. Illumination of the CPR enhances the efficiency of transmitting mechanosensory signals. It does so by increasing the SNR of mechanosensory input received from tailfan receptors. A sevenfold, nonlinear increase in the SNRs was observed with increasing light intensity, an effect especially pronounced for weak hydrodynamic stimuli. In comparison with the dark, illumination of the ganglion lowered the hydrodynamic threshold and heightened the response to suprathreshold stimulation. Unlike the SNR, the MDR is little affected by mechanosensory input. 4. These results are compared with simulated electronic activity from an analogue threshold model and are discussed with respect to the mechanism of stochastic resonance.

Neuropeptides in the human celiac/superior mesenteric ganglionic complex: An immunohistochemical study

The occurrence of vasoactive intestinal polypeptide (VIP), peptide histidine-isoleucine (PHI), calcitonin gene-related peptide (CGRP), substance P (SP), somatostatin (SOM), galanin (GAL) and enkephalins (ENK) is studied in the human celiac/superior mesenteric ganglionic complex of pre- and full-term new-borns, and adult subjects by means of immunohistochemistry. The antisera used labelled nerve fibres and terminal-like networks for each examined peptide, as well as VIP- and SOM-positive postganglionic neurons. Differences in the relative amount and density of the structures immunoreactive to the various peptides were observed. Moreover, variations in the amount and type of labelled elements were appreciable for each peptide when specimens from subjects at perinatal and adult ages were compared. Double-labelling immunofluorescence for SP and each other peptide showed that co-localization with SP is very frequent for CGRP, moderate to scarce for GAL and SOM, and rare to absent for PHI, VIP and ENK. VIP-, ENK- and CGRP-immunolabelled perikarya bearing the morphological features of the small intensely fluorescent (SIF) cells occurred in the organ. The presence of a paraganglion in one of the specimens examined allowed the detection of VIP- and ENK-positive cell bodies and VIP-, ENK-, SP- and GAL-like immunoreactive varicose nerve fibres in it. The results obtained provide substantial morphological data in support of the involvement of the examined peptides in the chemical interneuronal signalling in the human celiac/superior mesenteric ganglia.

Noradrenergic receptor function and aging: Beyond the binding

Quantitative studies of noradrenergic binding sites with aging probably underestimate the functional alterations in this, and other, central synaptic systems. Transmitters like norepinephrine, whose actions are based on amplification and interaction of interneuronal signalling capacity, rely on interactions among receptor subtypes for their complete effect. Thus, slight losses of one or both receptor subtypes with normal aging may have far more devastating effects.