Voltage-dependent Calcium Currents Research Articles

Relaxant Effect of Spermidine on Acethylcholine and High K+-induced Gastric Contractions of Guinea-Pig

In our previous study, we found that spermine and putrescine inhibited spontaneous and acetylcholine (ACh)-induced contractions of guinea-pig stomach via inhibition of L-type voltage-dependent calcium current (VDCC(L)). In this study, we also studied the effect of spermidine on mechanical contractions and calcium channel current (I(Ba)), and then compared its effects to those by spermine and putrescine. Spermidine inhibited spontaneous contraction of the gastric smooth muscle in a concentration-dependent manner (IC(50)=1.1+/-0.11 mM). Relationship between inhibition of contraction and calcium current by spermidine was studied using 50 mM high K(+)-induced contraction: Spermidine (5 mM) significantly reduced high K(+) (50 mM)-induced contraction to 37+/-4.7% of the control (p<0.05), and inhibitory effect of spermidine on I(Ba) was also observed at a wide range of test potential in current/voltage (I/V) relationship. Pre- and post-application of spermidine (5 mM) also significantly inhibited carbachol (CCh) and ACh-induced initial and phasic contractions. Finally, caffeine (10 mM)-induced contraction which is activated by Ca(2+)-induced Ca(2+) release (CICR),' was also inhibited by pretreatment of spermidine (5 mM). These findings suggest that spermidine inhibits spontaneous and CCh-induced contraction via inhibition of VDCC(L) and Ca(2+) releasing mechanism in guinea-pig stomach.

Serotonin Modulates Dendritic Calcium Influx in Commissural Interneurons in the Mouse Spinal Locomotor Network

Commissural interneurons (CINs) help to coordinate left-right alternating bursting activity during fictive locomotion in the neonatal mouse spinal cord. Serotonin (5-HT) plays an active role in the induction of fictive locomotion in the isolated spinal cord, but the cellular targets and mechanisms of its actions are relatively unknown. We investigated the possible role of serotonin in modifying dendritic calcium currents, using a combination of two-photon microscopy and patch-clamp recordings, in identified CINs in the upper lumbar region. Dendritic calcium responses to applied somatic voltage-clamp steps were measured using fluorescent calcium indicator imaging. Serotonin evoked significant reductions in voltage-dependent dendritic calcium influx in about 40% of the dendritic sites studied, with no detectable effect in the remaining sites. We also detected differential effects of serotonin in different dendritic sites of the same neuron; serotonin could decrease voltage-sensitive calcium influx at one site, with no effect at a nearby site. Voltage-clamp studies confirmed that serotonin reduces the voltage-dependent calcium current in CINs. Current-clamp experiments showed that the serotonin-evoked decreases in dendritic calcium influx were coupled with increases in neuronal excitability; we discuss possible mechanisms by which these two seemingly opposing results can be reconciled. This research demonstrates that dendritic calcium currents are targets of serotonin modulation in a group of spinal interneurons that are components of the mouse locomotor network.

Coupling a change in intraluminal pressure to vascular smooth muscle depolarization: still stretching for an explanation

pressure-induced (myogenic) constriction of arterioles is believed to be a fundamental contributor to the local regulation of microvascular blood flow and pressure. Myogenic constriction provides a level of tone upon which vasodilators can act to lower resistance and match blood flow to metabolic

Voltage-Dependent Calcium Currents Are Enhanced in Nucleus of the Solitary Tract Neurons Isolated From Renal Wrap Hypertensive Rats

The nucleus of the solitary tract (NTS) is the central site of termination of baroreceptor afferents. We hypothesize that changes occur in voltage-gated calcium channels (VGCCs) within NTS neurons as a consequence of hypertension. Whole-cell patch-clamp recordings were obtained from adult normotensive (109+/-2 mm Hg; n=6 from 6 sham-operated and 31 nonsurgically treated) and hypertensive (158+/-6 mm Hg; n=24) rats. In some experiments, 4-(4-[dihexadecylamino]styryl)-N-methylpyridinium iodide was applied to the aortic nerve to visualize NTS neurons receiving baroreceptor synaptic contacts. Ba(2+) currents (500 ms; -80 mV prepotential; 500 ms voltage steps in 5-mV increments to +15mV) peaked between -20 and -10 mV and were blocked by 100 mum of Cd(2+). Peak VGCCs were not different comparing non-4-(4-[dihexadecylamino]styryl)-N-methylpyridinium iodide-labeled and 4-(4- [dihexadecylamino]styryl)-N-methylpyridinium iodide-labeled NTS neurons in hypertensive and normotensive rats. The peak VGCC was significantly greater in cells from hypertensive compared with normotensive rats for both non-DiA-labeled (P=0.02) and DiA-labeled (P=0.04) neurons. To separate high-voltage activated (HVA) and low-voltage activated (LVA) components of VGCCs, voltage ramps (-110 mV to +30 mV over 50 ms) were applied from a holding potential of -60 mV (LVA channels inactivated) and a holding potential of -100 mV (both LVA and HVA currents activated). HVA currents were subtracted from HVA+LVA currents to yield the LVA current. Peak LVA currents were not different between hypertensive (8.9+/-0.8 pA/pF) and normotensive (7.8+/-0.6 pA/pF) groups of NTS neurons (P=0.27). These results demonstrate that 4 weeks of renal wrap hypertension induce an increase in Ca(2+) influx through HVA VGCCs in NTS neurons receiving arterial baroreceptor inputs.

Effects of Polyamines on Contractility of Guinea-Pig Gastric Smooth Muscle

This study was designed to investigate the effects of polyamines on mechanical contraction and voltage-dependent calcium current (VDCC) of guinea-pig gastric smooth muscle. Mechanical contraction and calcium channel current (IBa) were recorded by isometric tension recording and whole-cell patch clamp technique. Spermine, spermidine and putrescine inhibited spontaneous contraction of the gastric smooth muscle in a concentration-dependent manner. Spermine (2 mM) reduced high K+ (50 mM)-induced contraction to 16±6.4% of the control (n=9), and significantly inhibited IBa in a reversible manner (p<0.05; IC50=0.8 mM). Pre- and post-treatment of tissue with spermine (2-5 mM, n=10) also inhibited acetylcholine (10 µM)-induced phasic contraction to 5±6.4% of the control. Inhibitory effect of spermine on IBa was observed at a wide range of test potentials of current/voltage (I/V) relationship (p<0.05), and steady-state activation of IBa was shifted to the right by spermine (p<0.05). Spermidine and putrescine (1 mM each) also inhibited IBa to 51±5.7% and 81±5.3% of the control, respectively. And putrescine (1 mM) inhibited IBa at whole tested potentials (p<0.05) without significant change of kinetics (p<0.05). Finally, 5 mM putrescine also inhibited high K+-induced contraction to 53±7.1% of the control (n=4). These findings suggest that polyamines inhibit contractions of guinea-pig gastric smooth muscle via inhibition of VDCC.

Adenosine Inhibits Activity of Hypocretin/Orexin Neurons by the A1 Receptor in the Lateral Hypothalamus: A Possible Sleep-Promoting Effect

Neurons in the lateral hypothalamus (LH) that contain hypocretin/orexin have been established as important promoters of arousal. Deficiencies in the hypocretin/orexin system lead to narcolepsy. The inhibition of hypocretin/orexin neurons by sleep-promoting neurotransmitters has been suggested as one part of the sleep regulation machinery. Adenosine has been identified as a sleep promoter and its role in sleep regulation in the basal forebrain has been well documented. However, the effect of adenosine on arousal-promoting hypocretin/orexin neurons has not been addressed, despite recent evidence that immunocytochemical visualization of adenosine receptors was detected in these neurons. In this study, we examined the hypothesis that adenosine inhibits the activity of hypocretin/orexin neurons by using electrophysiological methods in brain slices from mice expressing green fluorescent protein in hypocretin/orexin neurons. We found that adenosine significantly attenuated the frequency of action potentials without a change in membrane potential in hypocretin/orexin neurons. The adenosine-mediated inhibition arises from depression of excitatory synaptic transmission to hypocretin/orexin neurons because adenosine depresses the amplitude of evoked excitatory postsynaptic potential and the frequency of spontaneous and miniature excitatory postsynaptic currents in these neurons. At the cell body of the hypocretin/orexin neurons, adenosine inhibits voltage-dependent calcium currents without the induction of GIRK current. The inhibitory effect of adenosine is dose dependent, pertussis toxin sensitive, and mediated by A1 receptors. In summary, our data suggest that in addition to its effect in the basal forebrain, adenosine exerts its sleep-promoting effect in the LH by inhibition of hypocretin/orexin neurons.

Effects of hypoxia, anoxia, and metabolic inhibitors on KATP channels in rat femoral artery myocytes

Vascular ATP-sensitive potassium (KATP) channels have an important role in hypoxic vasodilation. Because KATP channel activity depends on intracellular nucleotide concentration, one hypothesis is that hypoxia activates channels by reducing cellular ATP production. However, this has not been rigorously tested. In this study we measured KATP current in response to hypoxia and modulators of cellular metabolism in single smooth muscle cells from the rat femoral artery by using the whole cell patch-clamp technique. KATP current was not activated by exposure of cells to hypoxic solutions (Po2 approximately 35 mmHg). In contrast, voltage-dependent calcium current and the depolarization-induced rise in intracellular calcium concentration ([Ca2+]i) was inhibited by hypoxia. Blocking mitochondrial ATP production by using the ATP synthase inhibitor oligomycin B (3 microM) did not activate current. Blocking glycolytic ATP production by using 2-deoxy-D-glucose (5 mM) also did not activate current. The protonophore carbonyl cyanide m-chlorophenylhydrazone (1 microM) depolarized the mitochondrial membrane potential and activated KATP current. This activation was reversed by oligomycin B, suggesting it occurred as a consequence of mitochondrial ATP consumption by ATP synthase working in reverse mode. Finally, anoxia induced by dithionite (0.5 mM) also depolarized the mitochondrial membrane potential and activated KATP current. Our data show that: 1) anoxia but not hypoxia activates KATP current in femoral artery myocytes; and 2) inhibition of cellular energy production is insufficient to activate KATP current and that energy consumption is required for current activation. These results suggest that vascular KATP channels are not activated during hypoxia via changes in cell metabolism. Furthermore, part of the relaxant effect of hypoxia on rat femoral artery may be mediated by changes in [Ca2+]i through modulation of calcium channel activity.

Pharmacological and molecular enhancement of learning in aging and Alzheimer’s disease

When animals learn hippocampus-dependent associative and spatial tasks such as trace eyeblink conditioning and the water maze, CA1 hippocampal neurons become more excitable as a result of reductions in the post-burst, slow afterhyperpolarization. The calcium-activated potassium current that mediates this afterhyperpolarization is activated by the calcium influx that occurs when a series of action potentials fire and serves as a modulator of neuronal firing frequency. As a result, spike frequency accommodation is also reduced after learning. Neuronal calcium buffering processes change and/or voltage-dependent calcium currents increase during aging; leading to enhancements in the slow afterhyperpolarization, increased spike frequency accommodation and age-associated impairments in learning. We describe a series of studies done to characterize this learning-specific enhancement in intrinsic neuronal excitability and its converse in aging brain. We have also combined behavioral pharmacology and biophysics in experiments demonstrating that compounds that increase neuronal excitability in CA1 pyramidal neurons also enhance learning rate of hippocampus-dependent tasks, especially in aging animals. The studies reviewed here include those using nimodipine, an L-type calcium current blocker that tends to cross the blood-brain barrier; metrifonate, a cholinesterase inhibitor; CI1017, a muscarinic cholinergic agonist; and galantamine, a combined cholinesterase inhibitor and nicotinic agonist. Since aging is the chief risk factor for Alzheimer’s disease, a disease that targets the hippocampus and associated brain regions and markedly impairs hippocampus-dependent learning, these compounds have potential use as treatments for this disease. Galantamine has been approved by the USDA for this purpose. Finally, we have extended our studies to the TG2576 transgenic mouse model of Alzheimer’s disease (AD), that overproduces amyloid precursor protein (APP) and increases levels of toxic β-amyloid in the brain. Not only do these mice show deficits in hippocampus-dependent learning as they age, but their hippocampal neurons show a reduced capacity to increase their levels of intrinsic excitability with reductions in the slow afterhyperpolarization after application of the muscarinic agonist carbachol. These TG2576 APP overproducing mice were crossed with BACE1 knockout mice, that do not produce β-amyloid because cleavage of APP by the β-site APP cleaving enzyme 1 (BACE1) is a critical step in its formation. Not only was hippocampus-dependent learning rescued in the bigenic TG2576-BACE1 mice, but the capacity of hippocampal neurons to show normal enhancements of intrinsic excitability was restored. The series of studies reviewed here support our hypothesis that enhancement in intrinsic excitability by reductions in calcium-activated potassium currents in hippocampal neurons is an important cellular mechanism for hippocampus-dependent learning.

Modulation of L-type calcium current in rat cardiac myocytes by sulfur dioxide derivatives

The effects of sulfur dioxide (SO 2) derivatives (bisulfite and sulfite, 1:3 M/M) on voltage-dependent L-type calcium current ( I Ca,L) in isolated rat ventricular myocytes were studied using the whole cell patch-clamp technique. SO 2 derivatives increased I Ca,L in a concentration-dependent manner. SO 2 derivatives shifted both the steady-state activation and the inactivation curves of I Ca,L to more positive potentials, the effect on the latter being more pronounced. SO 2 derivatives markedly accelerated the recovery of I Ca,L from inactivation. SO 2 derivatives also significantly shortened the fast and slow time constants of inactivation. These results suggested that SO 2 inhalation might cause cardiac myocyte injury through increasing intracellular calcium via voltage-gated calcium channels.

Ca2+ currents and voltage responses in Type I and Type II hair cells of the chick embryo semicircular canal

Type I and Type II hair cells, and Type II hair cells located in different zones of the semicircular canal crista, express different patterns of voltage-dependent K channels, each one specifically shaping the hair cell receptor potential. We report here that, close to hatching, chicken embryo semicircular canal Type I and Type II hair cells express a similar voltage-dependent L-type calcium current (I(Ca)), whose main features are: activation above -60 mV, fast activation kinetics, and scarce inactivation. I(Ca) should be already active at rest in Zone 1 Type II hair cells, whose resting membrane potential was on average slightly less negative than -60 mV. Conversely, I(Ca) would not be active at rest in Type II hair cells from Zone 2 and 3, nor in Type I hair cells, since their resting membrane potential was significantly more negative than -60 mV. However, even small depolarising currents would activate I(Ca) steadily in Zone 2 and 3 Type II hair cells, but not in Type I hair cells because of the robust repolarising action of their specific array of K(+) currents. The implications of the present findings in the afferent discharge are discussed.

Serum-free culture of rat postnatal neurons derived from the dorsal motor nucleus of the vagus

Previous studies on dorsal motor nucleus of the vagus (DMNV) neurons have mainly used in vivo animal models and in vitro brainstem slices. Primary culture of postnatal DMNV neurons in defined serum free medium has not been reported. We report a method for culture of postnatal rat DMNV neurons using serum free medium. Cultured DMNV neurons contain both Hu positive precursor cells and mature cells staining positively for microtubule associated protein 2 (MAP2) and choline acetyltransferase. Exposure of cultured DMNV neurons to glutamate (10 −7 to 10 −3 M) induced an increase in intracellular calcium concentration ([Ca 2+] i ) in a dose-dependent manner, indicating the functional presence of glutamate receptors. Voltage-dependent calcium currents were present in cultured DMNV neurons. Active cell proliferation was demonstrated by BrdU incorporation. Upon removal of βFGF, the percentage of MAP2 positive mature neurons was significantly increased from 36 ± 3 to 73 ± 3%. Our study demonstrates that postnatal rat DMNV neurons cultured in serum free medium retain morphological and physiological characteristics of DMNV neurons in situ.

Action of aluminum on high voltage-dependent calcium current and its modulation by ginkgolide B

To investigate the effect of aluminum (Al) on high voltage-dependent calcium current (I(HVA)) and its modulation by ginkgolide B (Gin B). The whole-cell, patch-clamp technique was used to record IHVA from acutely isolated hippocampal CA1 pyramydal neurons in rats. Al 0.1 mmol/L (low concentration) reduced I(HVA); Al 0.75 and 1.0 mmol/L (high concentrations) increased I(HVA), and Al decreased and increased I(HVA) at intermediate concentrations of 0.25 and 0.5 mmol/L. The increase of I(HVA) by Al 1.0 mmol/L was enhanced by the adenylyl cyclase (AC) agonist forskolin and was partly abolished by the cyclic adenosine monophosphate (cAMP)-dependent protein kinase A (PKA) antagonist H-89, whereas the decrease observed with Al 0.1 mmol/L was neither reversed by forskolin nor affected by H-89. Gin B had no effect on I(HVA) in normal neurons, but canceled the increase in I(HVA) by 1.0 mmol/L Al. The results indicate that the mechanism of Al affecting I(HVA) differs at different concentrations, and this may be attributed to its complex actions. Gin B could prevent neurons from injury by inhibiting calcium influx.

Neuropeptide Y inhibits hypocretin/orexin neurons by multiple presynaptic and postsynaptic mechanisms: tonic depression of the hypothalamic arousal system.

Neurons that release neuropeptide Y (NPY) have important effects on hypothalamic homeostatic regulation, including energy homeostasis, and innervate hypocretin neurons. Using whole-cell patch-clamp recording, we explored NPY actions on hypocretin cells identified by selective green fluorescent protein expression in mouse hypothalamic slices. NPY reduced spike frequency and hyperpolarized the membrane potential of hypocretin neurons. The NPY hyperpolarizing action persisted in tetrodotoxin (TTX), was mimicked by Y1 receptor-selective agonists [Pro34]-NPY and [D-Arg25]-NPY, and was abolished by the Y1-specific antagonist BIBP3226 [(R)-N2-(diphenylacetyl)-N-[(4-hydroxyphenyl)methyl]-D-arginine-amide], consistent with a direct activation of postsynaptic Y1 receptors. NPY induced a current that was dependent on extracellular potassium, reversed near the potassium equilibrium potential, showed inward rectification, was blocked by extracellular barium, and was abolished by GDP-betaS in the recording pipette, consistent with a G-protein-activated inwardly rectifying K+ (GIRK) current. [Pro34]-NPY evoked, and BIBP3226 blocked, the activation of the GIRK-type current, indicating mediation by a Y1 receptor. NPY attenuated voltage-dependent calcium currents mainly via a Y1 receptor subtype. BIBP3226 increased spontaneous spike frequency, suggesting an ongoing Y1 receptor-mediated NPY inhibition. In TTX, miniature EPSCs were reduced in frequency but not amplitude by NPY, NPY13-36, and [D-Trp32]-NPY, but not by [Pro34]-NPY, suggesting the presynaptic inhibition was mediated by a Y2/Y5 receptor. NPY had little effect on GABA-mediated miniature IPSCs but depressed spontaneous IPSCs. Together, these data support the view that NPY reduces the activity of hypocretin neurons by multiple presynaptic and postsynaptic mechanisms and suggest NPY axons innervating hypocretin neurons may tonically attenuate hypocretin-regulated arousal.

Estimating use-dependent synaptic gain in autonomic ganglia by computational simulation and dynamic-clamp analysis.

Biological gain mechanisms regulate the sensitivity and dynamics of signaling pathways at the systemic, cellular, and molecular levels. In the sympathetic nervous system, gain in sensory-motor feedback loops is essential for homeostatic regulation of blood pressure and body temperature. This study shows how synaptic convergence and plasticity can interact to generate synaptic gain in autonomic ganglia and thereby enhance homeostatic control. Using a conductance-based computational model of an idealized sympathetic neuron, we simulated the postganglionic response to noisy patterns of presynaptic activity and found that a threefold amplification in postsynaptic spike output can arise in ganglia, depending on the number and strength of nicotinic synapses, the presynaptic firing rate, the extent of presynaptic facilitation, and the expression of muscarinic and peptidergic excitation. The simulations also showed that postsynaptic refractory periods serve to limit synaptic gain and alter postsynaptic spike timing. Synaptic gain was measured by stimulating dissociated bullfrog sympathetic neurons with 1-10 virtual synapses using a dynamic clamp. As in simulations, the threshold synaptic conductance for nicotinic excitation of firing was typically 10-15 nS, and synaptic gain increased with higher levels of nicotinic convergence. Unlike the model, gain in neurons sometimes declined during stimulation. This postsynaptic effect was partially blocked by 10 microM Cd2+, which inhibits voltage-dependent calcium currents. These results support a general model in which the circuit variations observed in parasympathetic and sympathetic ganglia, as well as other neural relays, can enable functional subsets of neurons to behave either as 1:1 relays, variable amplifiers, or switches.

Effect of dendritic location and different components of LTP expression on the bursting activity of hippocampal CA1 pyramidal cells

In recent in vivo experiments individual hippocampal CA1 pyramidal cells changed their bursting activity disperately in response to long-term potentiation (LTP) induced by stimulation in area CA3. We investigated that the LTP of which synapse system (CA3 recurrents or Schaffer collaterals) and which component of LTP at CA1 pyramidal cells (an increase in synaptic currents, a decrease in feed-forward inhibition, or an increase in voltage-dependent calcium currents) is responsible for these disperate effects. We found that only insertion of new calcium channels may lead to experimentally observed changes in bursting activity. In line with in vitro experimental findings, LTP in the basal dendrite resulted in a higher degree of potentiation than in the apical dendrite, which was accounted for by morphological differences between the two dendritic regions.

Differential regulation of neurotrophin expression in basal forebrain astrocytes by neuronal signals.

Nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin-3 (NT3) promote the function and/or survival of basal forebrain (BF) cholinergic neurons in vivo and in culture. The neurotrophin source is commonly thought to be targets of cholinergic neurons and the possibility that local glial sources support cholinergic neurons has not been well examined. These sources, however, may be critical to BF neurons before or even after they reach their targets. We investigated neurotrophin expression in BF astrocytes and its regulation by neural signals. Solution hybridization and immunocytochemical assays revealed that NGF, BDNF, and NT(3) mRNA and proteins were expressed in cultured BF astrocytes. To investigate roles of neuronal signals in neurotrophin regulation, effects of K(+), glutamate, and the cholinergic agonist carbachol were examined. These stimuli affected neurotrophin expression differentially. KCl increased BDNF mRNA but did not alter NGF or NT(3) mRNA. The effect was blocked by nifedipine, suggesting that it was mediated by L-type voltage-dependent calcium currents. Carbachol also increased BDNF mRNA levels without changing NGF or NT(3). Effects were blocked by the muscarinic antagonist, atropine. In contrast, glutamate increased both NGF and BDNF mRNA. NT(3) mRNA again was unaffected. The metabotropic agonist trans-(1S,3R)-1-amino-1,3-cyclopentanedicarboxylic acid (trans-ACPD) reproduced glutamate effects, whereas kainate or N-methyl-D-aspartate (NMDA) plus glycine did not. Lack of antagonism by ionotropic antagonists and blockade of glutamate effects by metabotropic antagonists confirmed metabotropic mediation. We suggest that BF astrocytes are local sources of neurotrophins for BF cholinergic neurons during development and are regulated differentially by specific neuronal signals. Critical neuronal-glial interactions may underlie basal forebrain function.

The Effect of Nickel and Manganese on Store-Operated Channels in Canine Pulmonary Arterial Smooth Muscle Cells

Background: Pulmonary vasoconstriction greatly contibutes to the elevated pulmonary vascular resistance in patients with pulmonary hypertension. A rise in cytosolic concentration in pulmonary arterial smooth muscle cells (PASMCs) is a major trigger for pulmonary vasoconstriction. Presently, little is known about the nature of the store-operated channels (SOCs) in PASMCs, even though store depletion-mediated capacitative entry (CCE) is a critical mechanism for refilling the empty sarcoplasmic reticulum (SR) with , and for maintaining a sustained increase in cyt. The goal of this in vitro study was to investigate the effects of nickel and manganese on store-operated channels in canine PASMCs. Methods: Isolated PASMCs were obtained from an enzymatically treated canine pulmonary artery. Currents were recorded at room temperature using the dialyzed whole cell recording technique. The protocol used to deplete the intracellular stores and to monitor the development of the store-operated currents, involved voltage-clamping cells at 0 mV to inactivate any voltage-dependent calcium currents. Crrents were recorded in response to a 200 ms voltage step from 120 to 40 mV in 20 mV increments every 15 seconds. Results: Simultaneous depletion of intracellular leads to a voltage-independent and linear store-operated current () reversal near 0 mV. Nickel and manganese inhibit . Conclusions: In canine PASMCs, the depletion of intracellular stores leads to the activation of Isoc, which is inhibited by nickel and manganese. These metals are non-specific inhibitors of non-selective cation channels. Our results indicate that - and -sensitive pathways may mediate entry, or that a class of non-selective cation channels may contribute to CCE in canine PASMCs.

PH changes in the invaginating synaptic cleft mediate feedback from horizontal cells to cone photoreceptors by modulating Ca2+ channels.

Feedback from horizontal cells (HCs) to cone photoreceptors plays a key role in the center-surround–receptive field organization of retinal neurons. Recordings from cone photoreceptors in newt retinal slices were obtained by the whole-cell patch-clamp technique, using a superfusate containing a GABA antagonist (100 μM picrotoxin). Surround illumination of the receptive field increased the voltage-dependent calcium current (ICa) in the cones, and shifted the activation voltage of ICa to negative voltages. External alkalinization also increased cone ICa and shifted its activation voltage toward negative voltages. Enrichment of the pH buffering capacity of the extracellular solution increased cone ICa, and blocked any additional increase in cone ICa by surround illumination. Hyperpolarization of the HCs by a glutamate receptor antagonist-augmented cone ICa, whereas depolarization of the HCs by kainate suppressed cone ICa. From these results, we propose the hypothesis that pH changes in the synaptic clefts, which are intimately related to the membrane voltage of the HCs, mediate the feedback from the HCs to cone photoreceptors. The feedback mediated by pH changes in the synaptic cleft may serve as an additional mechanism for the center-surround organization of the receptive field in the outer retina.

Amyotrophic lateral sclerosis patient IgG alters voltage dependence of Ca 2+ channels in dissociated rat motoneurons

Previous studies on sporadic amyotrophic lateral sclerosis (SALS) have detected IgG within motoneurons of patients and these IgGs have been shown to alter voltage dependent calcium channel activity in various cell types. The current study investigates whether IgG from categorized SALS patients alter voltage dependent calcium currents in rat motoneurons in culture. Patients were categorized based on onset and progression pattern. IgG yields were 38% higher in SALS patients compared to control subjects. Incubation with 1 mg/ml IgG from SALS patients did not cause visible toxicity, alter input resistance, capacitance or the maximal calcium conductance in rat motoneurons when compared to motoneurons incubated with control IgG. However, the activation curve of calcium current was shifted to the left in motoneurons treated with SALS IgG compared to control IgG.

Effects of acetylcholinesterase and butyrylcholinesterase on cell survival, neurite outgrowth, and voltage-dependent calcium currents of embryonic ventral mesencephalic neurons

The aim of this study was to investigate the effect of butyrylcholinesterase (BuChE) and acetylcholinesterase (AChE) on cell survival, neurite outgrowth and voltage-dependent calcium currents in developing rat ventral mesencephalic (VM) neurons. Both BuChE and AChE have been shown to promote neurite outgrowth in postnatnal preparations. However, the effect of these substances has never been investigated on rat embryonic VM cells, which are used in animal models of foetal transplantation as a treatment for Parkinson's disease. The effects of incubation with BuChE and tetrameric (G 4)- or monomeric (G 1)-AChE on cell survival and neurite outgrowth were characterised over a 7-day period on dopaminergic cells within embryonic VM cultures. The acute effects of these treatments on voltage-dependent calcium currents from embryonic VM cells were then investigated using whole-cell voltage-clamp recordings. The chronic effect of modulating voltage-dependent calcium channels was subsequently explored using the selective calcium channel antagonists ω-agatoxin IVA, ω-conotoxin GVIA, and nifedipine. The results presented here demonstrate firstly trophic effects of BuChE and G 4- and G 1-AChE upon dopaminergic neurite outgrowth, secondly that BuChE and G 4- and G 1-AChE have an inhibitory effect on voltage-dependent calcium currents, and finally that selective voltage-dependent calcium channel inhibitors also have trophic effects upon dopaminergic neurite outgrowth.