Voltage Sensitivity Of Activation Research Articles

Spinocerebellar ataxia type 6 knockin mice develop a progressive neuronal dysfunction with age-dependent accumulation of mutant Ca V 2.1 channels

Spinocerebellar ataxia type 6 (SCA6) is a neurodegenerative disorder caused by CAG repeat expansions within the voltage-gated calcium (Ca(V)) 2.1 channel gene. It remains controversial whether the mutation exerts neurotoxicity by changing the function of Ca(V)2.1 channel or through a gain-of-function mechanism associated with accumulation of the expanded polyglutamine protein. We generated three strains of knockin (KI) mice carrying normal, expanded, or hyperexpanded CAG repeat tracts in the Cacna1a locus. The mice expressing hyperexpanded polyglutamine (Sca6(84Q)) developed progressive motor impairment and aggregation of mutant Ca(V)2.1 channels. Electrophysiological analysis of cerebellar Purkinje cells revealed similar Ca(2+) channel current density among the three KI models. Neither voltage sensitivity of activation nor inactivation was altered in the Sca6(84Q) neurons, suggesting that expanded CAG repeat per se does not affect the intrinsic electrophysiological properties of the channels. The pathogenesis of SCA6 is apparently linked to an age-dependent process accompanied by accumulation of mutant Ca(V)2.1 channels.

Reduced voltage sensitivity of activation of P/Q-type Ca2+ channels is associated with the ataxic mouse mutation rolling Nagoya (tg(rol))

Reduced voltage sensitivity of activation of P/Q-type Ca2+ channels is associated with the ataxic mouse mutation rolling Nagoya (tg(rol))

The effects of catechol on various membrane conductances in lumbar sympathetic postganglionic neurones of the guinea-pig.

The effects of catechol on membrane properties in lumbar sympathetic postganglionic neurones isolated from guinea-pigs were studied in vitro in current and voltage clamp using single intracellular microelectrodes. Neurones with properties characteristic of two previously described classes of neurone (phasic and tonic) were studied. Catechol (3-12 mM) produced a few mV depolarization and a dose-dependent increase in membrane resistance which were both larger in tonic than in phasic neurones. In the presence of catechol, both phasic and tonic neurones fired only a single action potential at the beginning of a maintained depolarizing current step. In both neurone types, catechol reduced action potential amplitude and slowed its time course. The peak of the afterhyperpolarization became delayed and reduced in amplitude, particularly in tonic neurones. The time constant of inactivation of IA was reduced by catechol without change in the voltage sensitivity of activation or inactivation: IC50 was 3 mM in phasic and 4 mM in tonic neurones. Catechol also blocked a slow voltage-activated K+ current (resembling ID) that was present in many tonic neurones. Catechol did not modify the slow calcium-activated potassium current (gKCa1) or the anomalous rectifier; neither did it appear to affect the fast calcium-activated potassium current (IC) or the delayed rectifier. Catechol did not change the overall rate of spontaneous synaptic activity nor enhance the release of quanta of ACh from preganglionic terminals evoked by nerve stimulation. We conclude that, in addition to blocking IA, catechol blocks the slow ID-like current in sympathetic neurones. It also has a profound effect on the action potential probably by increasing inactivation of voltage-dependent Na+ channels. The change from tonic to phasic discharge in tonic neurones cannot be attributed solely to its effects on IA.

Effect of capsaicin and analogues on potassium and calcium currents and vanilloid receptors in Xenopus embryo spinal neurones.

1. The potassium current in embryo spinal neurones of Xenopus consists of at least two kinetically distinct components with overlapping voltage-dependencies of activation. We investigated whether capsaicin might specifically block these components in acutely dissociated neurones from stage 37/38 embryos by use of standard patch clamp techniques. 2. Capsaicin caused a time-dependent block of both the slow and fast components of the potassium current. The concentration-dependence was described by the Hill equation with a KD of 21 microM and a coefficient of 1.5 (n = 9-11 at each concentration). Differences between the observed and fitted values were not significant at the 5% level (chi(2) = 2.80, 6 degrees of freedom). 3. Capsaicin did not affect the time course or voltage-sensitivity of activation, but the steady-state block was voltage-dependent. The block could be relieved by hyperpolarization, and the rate of the removal of block was voltage- and time-dependent. The time constant for the blocking reaction was also voltage-dependent for voltage steps below +30 mV, but above this level it was voltage-independent. These results suggest that capsaicin blocks potassium channels by an open channel mechanism. 4. Other derivatives of vanillin, such as capsazepine, resiniferatoxin, and piperine also blocked potassium channels. Capsazepine and resiniferatoxin caused a greater block than similar concentrations of capsaicin, and in the case of capsazepine, the block was also clearly time-dependent. 5. Capsaicin and capsazepine also blocked calcium currents in a time-dependent manner. Fitting the Hill equation to the averaged data gave a KD of 43.5 microM, and a coefficient of 1.35 (n = 11 at each concentration). The fitted values were not significantly different from the observed means at the 5% level (chi(2) = 12.1, 6 degrees of freedom). 6. Six out of 29 Rohon-Beard sensory neurones responded to capsaicin with an inward current that appeared to be similar to the capsaicin activation of mammalian C sensory neurones. This response saturated at 10 microM capsaicin.

Modulation of a voltage-activated potassium channel by peptide growth factor receptors

Regulation of the activity of a cloned component of a voltage-activated K+ channel, Kv1.5, was studied by expressing the K+ channel and receptors for platelet-derived growth factor (PDGF) or fibroblast growth factor (FGF) simultaneously in Xenopus oocytes. Receptor activation mediated a decline in the Kv1.5 current amplitude, with a half-time of about 20 min. The reduction in K+ current amplitude occurred with little change in the kinetics or voltage sensitivity of activation. A similar phenomenon was found when the human thrombin or rat 5-HT1c receptors, two receptors that increase phospholipase C activity, were tested in coexpression experiments. A mutant FGF receptor, which does not activate phospholipase C-gamma 1 but retains several of its other functions, did not modulate the Kv1.5 current. Simultaneous injection of inositol trisphosphate and superfusion of phorbol 12-myristate 13-acetate reproduced the modulation of the Kv1.5 current. These results demonstrate that the PDGF and FGF receptors can modulate a voltage-activated K+ channel by increasing phospholipase C activity, and suggest that PDGF or FGF may be able to alter rapidly the electrical excitability of neurons.

Na+ channels in cardiac and neuronal cells derived from a mouse embryonal carcinoma cell line.

1. Cells from a pluripotent murine embryonal carcinoma cell line (P19) were differentiated in vitro into cells with neurone- and cardiac-like phenotypes. Cells treated with 0.5 microM retinoic acid developed into neurone-like cells possessing extensive neurites. Dimethyl sulphoxide treatment (0.5%) produced large, spontaneously contracting cell aggregates with many properties of cardiac cells. 2. The neurone- and cardiac-like cells contained voltage-sensitive Na+ channels with properties similar to those of native neuronal and cardiac cells. 3. We used whole-cell patch clamp techniques to measure inward currents from the neurone- and cardiac-like cells. Undifferentiated (untreated) cells had only small inward currents (peak of -0.15 nA in 150 mM external Na+). The peak inward current in the neurone-like and cardiac-like cells was -1.2 nA (in 154 mM external Na+) and -2.8 nA (in only 46 mM Na+), respectively. These large currents were absent when the external solution contained no Na+. 4. Tetrodotoxin (TTX) blocked the Na+ currents in the neurone- and cardiac-like cells in a dose-dependent manner. The Kd for TTX block of the Na+ current in the neurone-like cells was 6.7 nM. The Na+ current in the cardiac-like cells was much more resistant to TTX; the half-blocking concentration was two orders of magnitude higher, 710 nM. 5. The kinetic properties of the Na+ channel currents in the neurone- and cardiac-like cells were similar but developed over somewhat different voltage ranges. The voltage sensitivity of activation was similar in both cell types but the activation mid-point voltage was different: -12 mV in the neuronal cells and -34 mV for cardiac cells. Inactivation of the neuronal Na+ channels had a mid-point near -47 mV and was more sensitive to the membrane voltage than inactivation of the cardiac channels. The mid-point of inactivation for the cardiac Na+ channels was -80 mV.

Sodium currents during differentiation in a human neuroblastoma cell line.

The electrophysiological properties of a human neuroblastoma cell line, LA-N-5, were studied with the whole-cell configuration of the patch clamp technique before and after the induction of differentiation by retinoic acid, a vitamin A metabolite. Action potentials could be elicited from current clamped cells before the induction of differentiation, suggesting that some neuroblasts of the developing sympathetic nervous system are excitable. The action potential upstroke was carried by a sodium conductance, which was composed of two types of sodium currents, described by their sensitivity to tetrodotoxin (TTX) as TTX sensitive and TTX resistant. TTX-sensitive and TTX-resistant sodium currents were blocked by nanomolar and micromolar concentrations of TTX, respectively. The voltage sensitivity of activation and inactivation of TTX-resistant sodium current is shifted -10 to -30 mV relative to TTX-sensitive sodium current, suggesting that TTX-resistant sodium current could play a role in the initiation of action potentials. TTX-sensitive current comprised greater than 80% of the total sodium current in undifferentiated LA-N-5 cells. The surface density of total sodium current increased from 24.9 to 57.8 microA/microF after cells were induced to differentiate. The increase in total sodium current density was significant (P less than 0.05). The surface density of TTX-resistant sodium current did not change significantly during differentiation, from which we conclude that an increase in TTX-sensitive sodium current underlies the increase in total current.