Gap Junction Connections Research Articles

Gap junctions mediate large-scale Turing structures in a mean-field cortex driven by subcortical noise

One of the grand puzzles in neuroscience is establishing the link between cognition and the disparate patterns of spontaneous and task-induced brain activity that can be measured clinically using a wide range of detection modalities such as scalp electrodes and imaging tomography. High-level brain function is not a single-neuron property, yet emerges as a cooperative phenomenon of multiply-interacting populations of neurons. Therefore a fruitful modeling approach is to picture the cerebral cortex as a continuum characterized by parameters that have been averaged over a small volume of cortical tissue. Such mean-field cortical models have been used to investigate gross patterns of brain behavior such as anesthesia, the cycles of natural sleep, memory and erasure in slow-wave sleep, and epilepsy. There is persuasive and accumulating evidence that direct gap-junction connections between inhibitory neurons promote synchronous oscillatory behavior both locally and across distances of some centimeters, but, to date, continuum models have ignored gap-junction connectivity. In this paper we employ simple mean-field arguments to derive an expression for D2, the diffusive coupling strength arising from gap-junction connections between inhibitory neurons. Using recent neurophysiological measurements reported by Fukuda [J. Neurosci. 26, 3434 (2006)], we estimate an upper limit of D2 approximately 0.6cm2. We apply a linear stability analysis to a standard mean-field cortical model, augmented with gap-junction diffusion, and find this value for the diffusive coupling strength to be close to the critical value required to destabilize the homogeneous steady state. Computer simulations demonstrate that larger values of D2 cause the noise-driven model cortex to spontaneously crystalize into random mazelike Turing structures: centimeter-scale spatial patterns in which regions of high-firing activity are intermixed with regions of low-firing activity. These structures are consistent with the spatial variations in brain activity patterns detected with the BOLD (blood oxygen-level-dependent) signal detected with magnetic resonance imaging, and may provide a natural substrate for synchronous gamma-band rhythms observed across separated EEG (electroencephalogram) electrodes.

Dense gap-junction connections support dynamic Turing structures in the cortex

The recent report by Fukuda et al [1] provides convincing evidence for dense gap-junction connectivity between inhibitory neurons in the cat visual cortex, each neuron making 60 +/- 12 gap-junction dendritic connections with neurons in both the same and adjoining orientation columns. These resistive connections provide a source of diffusive current to the receiving neuron, supplementing the chemical-synaptic currents generated by incoming action-potential spike activity. Fukuda et al describe how the gap junctions form a dense and homogeneous electrical coupling of interneurons, and propose that this diffusion-coupled network provides the substrate for synchronization of neuronal populations. To date, large-scale population-based mathematical models of the cortex have ignored diffusive communication between neurons. Here we augment a well-established mean-field cortical model [2] by incorporating gap-junction-mediated diffusion currents, and we investigate the implications of strong diffusive coupling. The significant result is the model prediction that the 2D cortex can spontaneously generate centimetre-scale Turing structures (spatial patterns), in which regions of high-firing activity are intermixed with regions of low-firing activity (see Fig. ​Fig.1).1). Since coupling strength decreases with increases in firing rate, these patterns are expected to exchange contrast on a slow time-scale, with low-firing patches increasing their activity at the expense of high-firing patches. These theoretical predictions are consistent with the slowly fluctuating large-scale brain-activity images detected from the BOLD (blood oxygen-level-dependent) signal [3]. Figure 1 Diffusion-induced Turing patterns in a square cortex of side 25 cm. Panel a shows the case of zero diffusion: the cortex organizes into a diffuse, cloud-like pattern, but fails to generate a Turing structure. Panels b-d show increasing inhibitory diffusion. ...

White-noise susceptibility and critical slowing in neurons near spiking threshold

We present mathematical and simulation analyses of the below-threshold noisy response of two biophysically motivated models for excitable membrane due to H. R. Wilson: a squid axon ("resonator") and a human cortical neuron ("integrator"). When stimulated with a low-intensity white noise superimposed on a dc control current, both membrane types generate voltage fluctuations that exhibit critical slowing down--that is, the voltage responsiveness to noisy input currents grows in amplitude while slowing in frequency--as the membrane approaches spiking threshold from below. We define threshold unambiguously as that dc current that renders a zero real eigenvalue for the Jacobian matrix for the integrator neuron, and, for the resonator neuron, as the dc current that gives a complex eigenvalue pair whose real part is zero. Using a linear Ornstein-Uhlenbeck analysis, we give exact small-noise expressions for the variance, power spectrum, and correlation function of the voltage fluctuations, and we derive the scaling laws for the divergence of susceptibility and correlation times for approach to threshold. We compare these predictions with numerical simulations of the nonlinear stochastic equations, and demonstrate that, provided the white-noise perturbations are kept sufficiently small, the linearized theory works well. These predictions should be testable in the laboratory using a current-clamped cell configuration. If confirmed, then the proximity of a neuron to its spike-transition point can be judged by measuring its subthreshold susceptibility to white-noise stimulation. We postulate that such temporally correlated fluctuations could provide a means of subthreshold signaling via gap-junction connections with neighboring neurons.

Hypoxia disrupts intestinal gap junctions through activation of a PP1-dependent phosphatase and the de-phosphorylation of connexin43 in the pathogenesis of necrotizing enterocolitis

Introduction: Necrotizing enterocolitis (NEC) develops after a period of global hypoxia, and is characterized by a disruption in the intestinal barrier. The mechanisms by which hypoxia leads to barrier dysfunction remain largely unknown. We have shown that intestinal barrier function requires gap junctions comprised of connexin43 (Cx43) that must be phosphorylated to function. We now hypothesize that hypoxia leads to an impairment in intestinal gap junction function in vitro and in vivo, and sought to determine the mechanisms involved. Methods: IEC-6 cells were subjected to hypoxia in a flow regulated, vacuum sealed hypoxia chamber (1% O2, 37°C) for 1-12h. The expression and phosphorylation of Cx43 were assessed by SDS-PAGE and confocal immunofluorescence. To induce NEC, newborn rats were subjected to global hypoxia (10mins 5% O2) and formula gavage, and mucosal scrapings were obtained from the terminal ileum and subjected to SDS-PAGE. To restore Cx43 phosphorylation, IEC-6 cells were pre-treated with a variety of tyrosine phosphatase inhibitors, including pervanadate(10uM,1h) and calyculin A(2nM,30nM). Results: Hypoxic treatment caused a time-dependent decrease in the phosphorylation of Cx43, which correlated with a decrease in gap junction communication between adjacent enterocytes. There were no effects of hypoxia on the expression of total Cx43, or on cell viability. To verify the in vivo significance of these findings, phosphorylation of Cx43 was also significantly decreased in ileal mucosal scrapings from newborn rats subjected to global hypoxia and formula gavage as compared to control, breast fed animals. Strikingly, pre-treatment of IEC-6 cells with the protein phosphatase 1 (PP1) dependent phosphatase inhibitor calyculin A led to the restoration of phosphorylation of Cx43 and enhanced gap junction connectivity, indicating that a hypoxia-induced phosphatase mediated the effect. Conclusion: Hypoxia inhibits gap junction function in enterocytes through the activation of a protein phosphatase 1 dependent phosphatase resulting in the de-phosphorylation of Cx43. These findings suggest that further targeting of this hypoxia-induced phosphatase may provide a novel therapeutic strategy in the management of diseases of intestinal inflammation like NEC.

STRUCTURAL AND FUNCTIONAL PROPERTIES OF HOMOLOGOUS ELECTRICAL SYNAPSES BETWEEN RETINAL AMACRINE CELLS

Retinal amacrine cells regulate activities of retinal ganglion cells, the output neurons to higher visual centers, through cellular mechanism of lateral inhibition in the inner plexiform layer (IPL). Electrical properties of gap junction networks between amacrine cells in the IPL were investigated using combined techniques of intracellular recordings, Lucifer yellow and Neurobiotin injection, dual patch-clamp recordings and high voltage electron microscopy in isolated retinas of cyprinid fish. Six types of gap-junctionally connected amacrine cells were classified after their light-evoked responses to light flashes were recorded. Among them, gap junction networks of three types of amacrine cells were studied with structure-function correlation analysis. Cellular morphology of intercellular connections between three homologous cell classes was characterized. The interconnections between laterally extending dendrites in the IPL were localized at dendritic tip terminals. Three types of cells presented the dendrodendritic connections of tip-contact manner in the homologous cell population. High voltage as well as conventional electron microscopy revealed gap junctions between the dendritic tips of Neurobiotin-coupled cells. Receptive field properties of these amacrine cells were examined, displacing a slit of light along the distance from recording sites in the dorsal intermediate region of the retina. Receptive field size, space length constant, response latency and conduction velocity were measured. Spatial and temporal properties of receptive fields were symmetric along horizontally expanding dendrites in the dorsal retina. Simultaneous dual patch-clamp recordings revealed that the lateral gap junction connections between homologous amacrine cells expressed bidirectional electrical synapses passing Na(+) spikes. These results demonstrate that bidirectional electrical transmission in gap junction networks of these amacrine cells is symmetric along the lateral gap junction connections between horizontally extending dendrites. Lateral inhibition regulated by amacrine cells in the IPL appears to be associated with the directional extension of the dendrites and the orientation of dendrodendritic gap junctions.

Regeneration of dystrophin-expressing myocytes in the mdx heart by skeletal muscle stem cells

Cell transplantation holds promise as a potential treatment for cardiac dysfunction. Our group has isolated populations of murine skeletal muscle-derived stem cells (MDSCs) that exhibit stem cell-like properties. Here, we investigated the fate of MDSCs after transplantation into the hearts of dystrophin-deficient mdx mice, which model Duchenne muscular dystrophy (DMD). Transplanted MDSCs generated large grafts consisting primarily of numerous dystrophin-positive myocytes and, to a lesser degree, dystrophin-negative non-myocytes that expressed an endothelial phenotype. Most of the dystrophin-positive myocytes expressed a skeletal muscle phenotype and did not express a cardiac phenotype. However, some donor myocytes, located at the graft-host myocardium border, were observed to express cardiac-specific markers. More than half of these donor cells that exhibited a cardiac phenotype still maintained a skeletal muscle phenotype, demonstrating a hybrid state. Sex-mismatched donors and hosts revealed that many donor-derived cells that acquired a cardiac phenotype did so through fusion with host cardiomyocytes. Connexin43 gap junctions were not expressed by donor-derived myocytes in the graft. Scar tissue formation in the border region may inhibit the fusion and gap junction connections between donor and host cells. This study demonstrates that MDSC transplantation warrants further investigation as a potential therapy for cardiac dysfunction in DMD.

The Anesthetic Cascade

The Anesthetic Cascade

Further characterization of muscarinic agonist-induced epileptiform bursting activity in immature rat piriform cortex, in vitro

The characteristics of muscarinic acetylcholine receptor agonist-induced epileptiform bursting seen in immature rat piriform cortex slices in vitro were further investigated using intracellular recording, with particular focus on its postnatal age-dependence (P+14–P+30), pharmacology, site(s) of origin and the likely contribution of the muscarinic acetylcholine receptor agonist-induced post-stimulus slow afterdepolarization and gap junction functionality toward its generation. The muscarinic agonist, oxotremorine-M (10μM), induced rhythmic bursting only in immature piriform cortex slices; however, paroxysmal depolarizing shift amplitude, burst duration and burst incidence were inversely related to postnatal age. No significant age-dependent changes in neuronal membrane properties or postsynaptic muscarinic responsiveness accounted for this decline. Burst incidence was higher when recorded in anterior and posterior regions of the immature piriform cortex. In adult and immature neurones, oxotremorine-M effects were abolished by M1-, but not M2-muscarinic acetylcholine receptor-selective antagonists. Rostrocaudal lesions, between piriform cortex layers I and II, or layer III and endopiriform nucleus in adult or immature slices did not influence oxotremorine-M effects; however, the slow afterdepolarization in adult (but not immature) lesioned slices was abolished. Gap junction blockers (carbenoxolone or octanol) disrupted muscarinic bursting and diminished the slow afterdepolarization in immature slices, suggesting that gap junction connectivity was important for bursting. Our data show that neural networks within layers II–III function as primary oscillatory circuits for burst initiation in immature rat piriform cortex during persistent muscarinic receptor activation. Furthermore, we propose that muscarinic slow afterdepolarization induction and gap junction communication could contribute towards the increased epileptiform susceptibility of this brain area.

Connexin43, the major gap junction protein of astrocytes, is down-regulated in inflamed white matter in an animal model of multiple sclerosis

Both multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE), its animal model, involve inflammatory attack on central nervous system (CNS) white matter, leading to demyelination and axonal damage. Changes in astrocytic morphology and function are also prominent features of MS and EAE. Resting astrocytes form a network that is interconnected through gap junctions, composed mainly of connexin43 (Cx43) protein. Although astrocytic gap junctional connectivity is known to be altered in many CNS pathologies, little is known about Cx43 expression in inflammatory demyelinating disease. Therefore, we evaluated the expression of Cx43 in spinal cords of EAE mice compared with healthy controls. Lumbar ventral white matter areas were heavily infiltrated with CD11beta-immunoreactive monocytes, and within these infiltrated regions loss of Cx43 immunoreactivity was evident. These regions also showed axonal dystrophy, demonstrated by the abnormally dephosphorylated heavy-chain neurofilament proteins. Astrocytes in these Cx43-depleted lesions were strongly glial fibrillary acidic protein reactive. Significant loss (38%) of Cx43 protein in EAE mouse at the lumbar portion of spinal cords was confirmed by Western blot analysis. Decreased Cx43 transcript level was also observed on cDNA microarray analysis. In addition to changes in Cx43 expression, numerous other genes were altered, including those encoding adhesion and extracellular matrix proteins. Our data support the notion that, in addition to damage of myelinating glia, altered astrocyte connectivity is a prominent feature of inflammatory demyelination.

Dendrodendritic Electrical Synapses between Mammalian Retinal Ganglion Cells

Electrical synapses between alpha-type ganglion cells were detected using combined techniques of dual patch-clamp recordings, intracellular labeling, electron microscopy, and channel subunit connexin immunocytochemistry in the albino rat retina. After intracellular injection of Neurobiotin into alpha-cells of inner (ON-center) and outer (OFF-center) ramifying types, measurement of tracer coupling resulted in a preferentially homologous occurrence among cells of the same morphological type (n = 19 of 24). In high-voltage as well as conventional electron microscopic analysis, direct dendrodendritic gap junctions (average size, 0.86 mum long) were present in contact sites between tracer-coupled alpha-cells. In simultaneous dual whole-cell recordings from pairs of neighboring alpha-cells, these cells generated TTX-sensitive sustained spiking against extrinsic current injection, and bidirectional electrical synapses (maximum coupling coefficient, 0.32) with symmetrical junction conductance (average, 1.35 nS) were observed in pairs with cells of the same morphological type. Precise temporal synchronization of spike activity (average time delay, 2.7 msec) was detected when depolarizing currents were simultaneously injected into the pairs. To address whether physiologically identified electrical synapses constitute gap junctional connectivity between cell pairs, identified neuronal connexin36 immunoreactivity was undertaken in Lucifer yellow-labeled cell pairs after patch-clamp recordings. All alpha-cells expressed connexin36, and confocal laser-scanning imaging demonstrated that connexin36 is primarily located at dendritic crossings between electrically coupled cells (seven sites in a pair, on average). These results give conclusive evidence for electrical synapses via dendrodendritic gap junctions involving connexin36 in alpha retinal ganglion cells of the same physiological type.

Spinal gap junctions: Potential involvement in pain facilitation

Abstract Glia are now recognized as important contributors in pathological pain creation and maintenance. Spinal cord glia exhibit extensive gap junctional connectivity, raising the possibility that glia are involved in the contralateral spread of excitation resulting in mirror image pain. In the present experiments, the gap junction decoupler carbenoxolone was administered intrathecally after induction of neuropathic pain in response to sciatic nerve inflammation (sciatic inflammatory neuropathy) or partial nerve injury (chronic constriction injury). In both neuropathic pain models, a low dose of carbenoxolone reversed mirror image mechanical allodynia, while leaving ipsilateral mechanical allodynia unaffected. Ipsilateral thermal hyperalgesia was briefly attenuated. Critically, blockade of mechanical allodynia and thermal hyperalgesia was not observed in response to intrathecal glycyrrhizic acid, a compound similar to carbenoxolone in all respects but it does not decouple gap junctions. Thus, blockade of mechanical allodynia and thermal hyperalgesia by carbenoxolone does appear to reflect an effect on gap junctions. Examination of carbenoxolone's effects on intrathecal human immunodeficiency virus type 1 gp120 showed that blockade of pain facilitation might result, at least in part, via suppression of interleukin-1 and, in turn, interleukin-6. These data provide the first suggestion that spread of excitation via gap junctions might contribute importantly to inflammatory and traumatic neuropathic pain. Perspective The current studies provide evidence for involvement of gap junctions in spinal cord pain facilitation. Intrathecal carbenoxolone, a gap junction decoupler, reversed neuropathy-induced mirror image pain and intrathecal gp120-induced allodynia. In addition, it decreased gp120-induced proinflammatory cytokines. This suggests gap junction activation might lead to proinflammatory cytokine release by distantly activated glia.

Developmental changes in transmission of respiratory rhythm in the rat

We used cross-correlation to examine the short time-scale synchronisation of left and right phrenic nerve discharges in in-situ preparations of rats over a range of ages, to investigate the development of respiratory rhythm transmission to phrenic motoneurones. We found central peaks in the cross-correlograms, indicative of short time-scale synchronisation, at all ages (2-41 days), whose half-amplitude widths varied inversely with age (40-1.8 ms). In 10 preparations < or =5-days-old the central peaks were unaffected by a mid-sagittal section from C3 to C6. Carbenoxalone (CBX), a gap junction blocker, and its inactive analogue glycyrrhzic acid (GZA), eliminated central peaks in preparations younger than 12 days but not in older preparations. We concluded that in rats older than approximately 12 days short time-scale synchronisation is produced by bilaterally-projecting axons of medullary pre-motor neurones, whereas in younger rats it is due to pre-synaptic synchronisation of left and right medullary pre-motor neurones. While the latter mechanism may be gap junction connections, these experiments cannot unequivocally demonstrate it.

Defective Activation of c-Src in Cystic Fibrosis Airway Epithelial Cells Results in Loss of Tumor Necrosis Factor-α-induced Gap Junction Regulation

Tumor necrosis factor-alpha (TNF-alpha) signaling is central to the transmission of the innate immune response and subsequent activation of the adaptive immune system. The functioning of both systems is required for optimal clearance of pathogens from the airways. In cystic fibrosis (CF), dysfunction of the CF transmembrane conductance regulator (CFTR) is associated with recurrent pulmonary infections despite an intense inflammatory and immune response. We reported recently that TNF-alpha decreased gap junction connectivity in non-CF airway cells, a mechanism that was absent in CF cells expressing the DeltaPhe-508 mutant of CFTR. We have now identified the tyrosine kinase c-Src as a possible pathway between the mediators of inflammation and the gap junction protein connexin43 (Cx43). Indeed, TNF-alpha increased the proportion of activated c-Src in non-CF airway cells. Moreover, pharmacological antagonists and expression in non-CF cells of a dominant negative construct of c-Src prevented Cx43 channel closure by TNF-alpha. Finally, gap junction channel closure was prevented by expression of a Cx43 mutant lacking tyrosine phosphorylation sites for c-Src. Additional experiments showed that activation of c-Src was defective in CF airway cells but rescued in CFTR-corrected CF cells. These data suggest that CFTR dysfunction is associated with altered TNF-alpha signaling, resulting in the persistence of gap junction connectivity in CF airway cells. We propose that altered regulation of c-Src may contribute to the dysregulated inflammatory response that is characteristic of the CF phenotype.

Neighborly interactions of metabolically-activated astrocytes in vivo

Metabolic responses of brain cells to a stimulus are governed, in part, by their enzymatic specialization and interrelationships with neighboring cells, and local shifts in functional metabolism during brain activation are likely to be influenced by the neurotransmitter system, subcellular compartmentation, and anatomical structure. Selected examples of functional activation illustrate the complexity of metabolic interactions in working brain and of interpretation of changes in brain lactate levels. The major focus of this article is the disproportionately higher metabolism of glucose compared to oxygen in normoxic brain, a phenomenon that occurs during activation in humans and animals. The glucose utilized in excess of oxygen is not fully explained by accumulation of glucose, lactate, or glycogen in brain or by lactate efflux from brain to blood. Thus, any lactate derived from the excess glucose could not have been stoichiometrically exported to and metabolized by neighboring neurons because oxygen consumption would have otherwise increased and matched that of glucose. Metabolic labeling of tricarboxylic acid cycle-derived amino acids increased during brief sensory stimulation, reflecting a rise in oxidative metabolism. Brain glycogen is mainly in astrocytes, and its level falls throughout the stimulus and early post-activation interval. Glycogenolysis cannot be accounted for by lactate accumulation or oxidation; there must be rapid product clearance. Glycogen restoration is slow and diversion of glucose from oxidative pathways for its re-synthesis could reduce the global O2/glucose uptake ratio; astrocytes could downshift this ratio for up to an hour after 5min stimulus. Morphological studies of astrocytes reveal a paucity of cytoplasm and organelles in the fine processes that surround synapses and form gap junction connections with neighboring astrocytes. Specialized regions of astrocytes, e.g. their endfeet and thin peripheral lamellae, are likely to have compartmentalized metabolic activities. Anatomical constraints imposed upon the fine processes might require preferential utilization of glycolysis to satisfy their energy demands, but rapid lactate clearance would then be essential, since its accumulation would inhibit glycolysis. Gap junctional connections between neighboring astrocytes provide a mechanism for rapid metabolite spreading via the astrocytic syncytium and elimination of by-products. Local structure–function relationships need to be incorporated into experimental models of neuron–astrocyte and astrocyte–astrocyte interactions in working brain.

Src signaling links mediators of inflammation to Cx43 gap junction channels in primary and transformed CFTR-expressing airway cells.

Dysfunction of the cystic fibrosis transmembrane conductance regulator (CFTR) is associated with recurrent pulmonary infections and inflammation. We previously reported that tumor necrosis factor (TNF)-alpha decreases gap junction connectivity in cell lines derived from the airway epithelium of non-cystic fibrosis (non-CF) subjects, a mechanism that was defective in cells derived from CF patients, and identified the tyrosine kinase c-Src as a possible bridge between TNF-alpha and Cx43. To examine whether this modulation also takes place in primary epithelial cells, the functional expression of Cx43 was studied in non-CF and CF airway cells, obtained from surgical polypectomies and turbinectomies, which were grown either on culture dishes or permeable filters. Expression of Cx43 was detected by immunofluorescence on cells grown under both culture conditions. Non-CF and CF airway cells also showed intercellular diffusion of Lucifer Yellow. Dye coupling was rapidly abolished in non-CF cells in the presence of TNF-alpha, lipopolysaccharide and lysophosphatidic acid, and could be prevented by tyrphostin47, an inhibitor of Src tyrosine kinases. This down-regulation, however, was not detected in CF airway cells. These data indicate that CFTR dysfunction is associated with altered Src signaling, resulting in the persistence of gap junction connectivity in primary and transformed CF airway cells.

Effects of nifedipine and nickel on plateau potentials generated in submucosal interstitial cells distributed in the mouse proximal colon.

The effects of nifedipine and nickel ions (Ni2+), known inhibitors of L- and T-type voltage-gated Ca-channels respectively, were investigated on plateau potentials recorded from submucosal interstitial cells distributed in the mouse proximal colon. Plateau potentials were generated at a frequency of about 15 times min(-1) and were formed of two components. The primary component had an initial fast rate of rise with a transient potential (rate of rise, 130 mV/s; peak amplitude, 35 mV) and was followed by a secondary plateau component with a sustained potential (amplitude, 25 mV; duration, 2.6 s). Each cell from which recordings were made was injected with neurobiotin. Subsequent morphological examination with a confocal microscope indicated successful visualization of injected cells only in the presence of 18beta-glycylrhetinic acid (an inhibitor of gap junctional connections), suggesting that these cells were dye-coupled with surrounding cells. The cells injected with neurobiotin exhibited an oval-shaped cell body with bipolar processes and were distributed in the submucosal layer, suggesting that they were submucosal interstitial cells of Cajal (ICC-SM). The plateau potentials were not altered by 0.01 microM nifedipine, but were reduced in duration by 0.1 microM nifedipine, and abolished by 1 microM nifedipine. The rate of rise of plateau potentials, but not their amplitude, was reduced by Ni2+ (> 10 microM), with no significant alteration of the membrane potential. In the presence of 100 microM Ni2+, the plateau potentials were changed to a triangular form. Thus, the plateau potentials were formed by two types of voltage-gated channel current: the initial component was produced by a Ni2+-sensitive channel current and the plateau component by a nifedipine-sensitive current. The possible involvement of two different types of voltage-gated Ca2+-channels in the generation of submucosal pacemaker potentials was discussed.

Increased incidence of gap junctional coupling between spinal motoneurones following transient blockade of NMDA receptors in neonatal rats.

Neonatal rat motoneurones are electrically coupled via gap junctions and the incidence of this coupling declines during postnatal development. The mechanisms involved in this developmental regulation of gap junctional communication are largely unknown. Here we have studied the role of NMDA receptor-mediated glutamatergic synaptic activity in the regulation of motoneurone coupling. Gap junctional coupling was demonstrated by the presence of graded, short latency depolarising potentials following ventral root stimulation, and by the transfer of the low molecular weight tracer Neurobiotin to neighbouring motoneurones. Sites of close apposition between the somata and/or dendrites of the dye-coupled motoneurones were identified as potential sites of gap junctional coupling. Early postnatal blockade of the NMDA subtype of glutamate receptors using the non-competitive antagonist dizocilpine maleate (MK801) arrested the developmental decrease in electrotonic and dye coupling during the first postnatal week. These results suggest that the postnatal increase in glutamatergic synaptic activity associated with the onset of locomotion promote the loss of gap junctional connections between developing motoneurones.

β and γ Frequency Synchronization by Dendritic GABAergic Synapses and Gap Junctions in a Network of Cortical Interneurons

Distinct interneuron populations innervate perisomatic and dendritic regions of cortical cells. Perisomatically terminating GABAergic inputs are effective in timing postsynaptic action potentials, and basket cells synchronize each other via gap junctions combined with neighboring GABAergic synapses. The function of dendritic GABAergic synapses in cortical rhythmicity, and their interaction with electrical synapses is not understood. Using multiple whole-cell recordings in layers 2-3 of rat somatosensory cortex combined with light and electron microscopic determination of sites of interaction, we studied the interactions between regular spiking nonpyramidal cells (RSNPCs). Random samples of unlabeled postsynaptic targets showed that RSNPCs placed GABAergic synapses onto dendritic spines (53 +/- 12%) and shafts (45 +/- 10%) and occasionally somata (2 +/- 4%). GABAergic interactions between RSNPCs were mediated by 4 +/- 2 axodendritic synapses and phased postsynaptic activity at beta frequency but were ineffective in phasing at gamma rhythm. Electrical interactions of RSNPCs were transmitted via two to eight gap junctions between dendritic shafts and/or spines. Elicited at beta and gamma frequencies, gap junctional potentials timed postsynaptic spikes with a phase lag, however strong electrical coupling could synchronize presynaptic and postsynaptic activity. Combined unitary GABAergic and gap junctional connections of moderate strength produced beta and gamma frequency synchronization of the coupled RSNPCs. Our results provide evidence that dendritic GABAergic and/or gap junctional mechanisms effectively transmit suprathreshold information in a population of interneurons at behaviorally relevant frequencies. A coherent network of GABAergic cells targeting the dendrites could provide a pathway for rhythmic activity spatially segregated from perisomatic mechanisms of synchronization.

Cytokine Regulation of Gap Junction Connectivity: An Open-and-Shut Case or Changing Partners at the Nexus?

In this issue, Chanson et al 1 present data that document the failure of airway cells expressing the mutant form of the cystic fibrosis (CF) transmembrane conductance regulator (CFTR) gene to down-regulate gap junction connectivity after treatment with the cytokine tumor necrosis factor-α (TNF-α). However, this response could be restored by transfecting in the wild-type CFTR gene, suggesting that a functional CFTR gene is necessary for regulation of gap junctional communication by TNF-α. The authors postulate that the persistence of gap-junction connectivity in the inflamed CF airway epithelium permits the intercellular diffusion of signaling molecules that serve to activate neighboring cells, thus contributing to the excessive inflammatory response characteristic of the CF airway epithelium. Although these conclusions are based only on estimation of gap-junction connectivity using transfer of the gap-junction permeant dye Lucifer Yellow, the data fit well with several studies that have documented regulation of gap junction connectivity in a number of different cell types by proinflammatory cytokines such as TNF-α and interleukin-1β (IL-1β). 2-8 These interesting observations raise the questions of how inflammatory cytokines might regulate gap junction connectivity and in what way expression of the mutant CFTR gene could influence this response.

Dynamics of two electrically coupled chaotic neurons: experimental observations and model analysis.

Conductance-based models of neurons from the lobster stomatogastric ganglion (STG) have been developed to understand the observed chaotic behavior of individual STG neurons. These models identify an additional slow dynamical process calcium exchange and storage in the endoplasmic reticulum as a biologically plausible source for the observed chaos in the oscillations of these cells. In this paper we test these ideas further by exploring the dynamical behavior when two model neurons are coupled by electrical or gap junction connections. We compare in detail the model results to the laboratory measurements of electrically-coupled neurons that we reported earlier. The experiments on the biological neurons varied the strength of the effective coupling by applying a parallel, artificial synapse, which changed both the magnitude and polar-of the conductance between the neurons. We observed a sequence of bifarctions that took the neurons from strongly synchronized in-phase behavior. through uncorrelated chaotic oscillations to strongly synchronized and now regular out-of-phase behavior. The model calculations reproduce these observations quantitatively, indicating that slow subcellular processes could account for the mechanisms involved in the synchronization and regularization of the otherwise individual chaotic activities.