Ryanodine- and thapsigargin-insensitive Ca2+-induced Ca2+ release is primed by lowering external Ca2+ in rabbit autonomic neurons.

Rises in cytosolic Ca2+ induced by a high K+ concentration (30 or 60 mM) (K+-induced Ca2+ transient) were recorded by fluorimetry of Ca2+ indicators in cultured rabbit otic ganglion cells. When external Ca2+ ([Ca2+]o) was reduced to a micromolar (10-40 microM) or nanomolar (<10 nM) level prior to high-K+ treatment, K+-induced Ca2+ transients of considerable amplitude (50% of control) were generated in most cells, although those initiated at normal [Ca2+]o were reduced markedly or abolished by reducing [Ca2+]o during exposure to a high K+ concentration. Lowering [Ca2+]o alone occasionally caused a transient rise in cytosolic Ca2+. K+-induced Ca2+ transients at micromolar [Ca2+]o were repeatedly generated and propagated inwardly at a speed slower than that at normal [Ca2+]o, while those at nanomolar [Ca2+]o occurred only once. K+-induced Ca2+ transients at micromolar [Ca2+]o were not blocked by ryanodine (10 microM), carbonyl cyanide p-(trifluoromethoxy) phenylhydrazone (FCCP, 5 microM: at 20-22 degrees C but blocked at 31-34 degrees C) or thapsigargin (1-2 microM), but were blocked by Ni2+ (1 mM) or nicardipine (10 microM). Thus, there is a ryanodine-insensitive Ca2+-release mechanism in FCCP- and thapsigargin-insensitive Ca2+ stores in rabbit otic ganglion cells, which is primed by lowering [Ca2+]o and then activated by depolarization-induced Ca2+ entry. This Ca2+-induced Ca2+ release may operate when [Ca2+]o is decreased by intense neuronal activity.

Modes of propagation of Ca(2+)-induced Ca2+ release in bullfrog sympathetic ganglion cells.

How depolarization-induced Ca2+ entry or caffeine activates Ca(2+)-induced Ca2+ release (CICR) in the cytoplasm and nucleoplasm was studied by recording intracellular Ca2+ ([Ca2+]i) with a confocal microscope in cultured bullfrog sympathetic ganglion cells. The amplitude and propagation speed of voltage pulse-induced rises in [Ca2+]i were greater in the submembrane (< 5 microns depth) region than in the core region, and delayed and smaller, but significant, in the nucleus. Ryanodine and dantrolene reduced the rises in [Ca2+]i in both the cytoplasm and nucleus. A rapid application of high K+ solution induced global rises in [Ca2+]i in both the cytoplasm and nucleoplasm, which were decreased by dantrolene. Caffeine produced a slow, small rise in [Ca2+]i which grew into a global, regenerative rise both in the cytoplasm and nucleoplasm with some inward gradient in the cytoplasm. Each of the high [Ca2+]i phases during caffeine-induced [Ca2+]i oscillation began in the submembrane region, while low [Ca2+]i phases started in the core region. These results suggest that CICR activated by Ca2+ entry or caffeine occurs predominantly in the submembrane region causing an inwardly spreading Ca2+ wave or [Ca2+]i oscillations, and that the nuclear envelope can cause CICR in the nucleoplasm, which is delayed due to Ca2+ diffusion barrier at the nuclear pores.

Ca(2+)-induced Ca2+ release and its activation in response to a single action potential in rabbit otic ganglion cells.

1. Ryanodine-sensitive intracellular Ca2+ release activated by Ca2+ entry was studied with fura-2 fluorescence and intracellular voltage recording techniques in rabbit otic ganglion cells. 2. The removal of extracellular Ca2+ reduced sustained, transient or oscillatory rises in intracellular Ca2+ ([Ca2+]i) induced at high extracellular K+ and abolished the [Ca2+]i oscillation in cultured neurones. 3. Ryanodine (10 microM) transiently increased [Ca2+]i and reduced the amplitude and rate of rise of the high-K(+)-induced rise in [Ca2+]i, while caffeine (5 mM) produced a few transient rises in [Ca2+]i in most cultured cells and [Ca2+]i oscillation only in one cell. 4. The two components of the slow after-hyperpolarization (AHP) of an action potential in neurones of freshly isolated ganglia were dependent on extracellular Ca2+ and abolished by Ca2+ channel blockers, Cd2+ or Co2+. 5. The late component of AHP (LAHP), but not the initial component, in 'fresh' neurones increased in area with an increase in the preceding interval, was abolished by ryanodine (10 microM) and intracellularly injected EGTA, and mimicked by intracellular injection of Ca2+. 6. A ryanodine-sensitive Ca(2+)-induced Ca2+ release thus exists, operates in response to an action potential-induced Ca2+ entry and underlies LAHP in rabbit otic ganglion cells.

[Effects of byakushi and ogon on the hepatic drug metabolizing enzymes in rats].

The effect of single or daily oral administration of hot water extracts (HWE) from Byakushi or Ogon on rat hepatic drug-metabolizing enzymes were investigated in vivo. Enzymes were measured for 3 to 72 hr after single oral administration of HWE at the dose of 1.0 g/kg or 5.0 g/kg. Administration of 1.0 g/kg of Byakushi and 5.0 g/kg of Ogon inhibited aniline hydroxylase activity, while 5.0 g/kg of Byakushi inhibited it in the early phase, but increased it in the late phase. Byakushi inhibited aminopyrine N-demethylase activity, while 5.0 g/kg of Ogon increased it. Byakushi and Ogon decreased the amount of cytochrome P-450. Byakushi and Ogon increased the amount of cytochrome b5. Byakushi increased cytochrome c reductase activity 3 hr after administration and decreased it 6 and 12 hr after administration. In contrast, 1.0 g/kg of Ogon decreased cytochrome c reductase activity, and 5.0 g/kg increased it 6 hr after administration and decreased it 12 hr after administration. At 24 hr after the last administration to animals treated with a regimen of once a day administration of the HWE (0.1 or 1.0 g/kg) of Byakushi or Ogon for 14 days, the enzymes were measured. Byakushi decreased aminopyrine N-demethylase activity, the amount of cytochrome P-450, and cytochrome c reductase activity. Ogon decreased cytochrome c reductase activity. Byakushi altered the composition of cytochrome P-450 isozyme after daily administration.

Cyclic ADP-ribose modulates Ca2+ release channels for activation by physiological Ca2+ entry in bullfrog sympathetic neurons.

Although Ca(2+)-induced Ca2+ release (CICR) via ryanodine receptors has been found to occur in intact neurons, little is known about the physiological processes that regulate it. We studied the effects of cyclic ADP-ribose (cADPR) on CICR in cultured bullfrog sympathetic neurons by fura-2 fluorescence recording and patch-clamp techniques. cADPR applied through a patch pipette augmented action potential- or depolarizing pulse-induced rises in intracellular Ca2+ without a change in Ca2+ entry initiating the responses, but not in the presence of ryanodine. Likewise, cADPR enhanced a single or oscillatory rise(s) in intracellular Ca2+ induced by caffeine. These results strongly suggest that cADPR can be an endogenous modulator of ryanodine receptors in neurons.

Characteristics of Ca2+ release induced by Ca2+ influx in cultured bullfrog sympathetic neurones.

1. A rise in intracellular Ca2+ ([Ca2+]i) and a Ca2+ current (ICa) induced by a depolarizing pulse were simultaneously recorded by fura-2 or indo-1 fluorescence and whole-cell patch clamp techniques in cultured bullfrog sympathetic ganglion cells. 2. [Ca2+]i (calculated from the ratio of fura-2 fluorescences excited at 380 and 340 nm and recorded with a photomultiplier at > 492 nm) rose regeneratively (in most cells) during a command pulse (from -60 to 0 mV, 100 ms), continued to rise thereafter, peaked at 666 ms (on average) and decayed slowly with a half-decay time of 22.8 s. 3. Scanning a single horizontal line across the cytoplasm with an ultraviolet argon ion laser (351 nm) and recording indo-1 fluorescences at two wavelengths (peaked at 410 and 475 nm) with a confocal microscope demonstrated that [Ca2+]i beneath the cell membrane rose much faster than that in the deeper cytoplasm. The time course of the spatial integral of [Ca2+]i, however, corresponded well with that recorded with fura-2 fluorescence using a photomultiplier. 4. [Ca2+]i measured by fura-2 fluorescence ratio using a photomultiplier did not increase during a strong depolarizing pulse (-60 to +80 mV), but sometimes rose after the pulse. A depolarization-induced rise in [Ca2+]i ([Ca2+]i transient) was blocked in a Ca(2+)-free, EGTA solution, reduced by lowering the extracellular Ca2+ concentration ([Ca2+]o) to 0.45 or 0.9 mM and enhanced by raising [Ca2+]o to 7.2 or 14.4 nM. 5. The extracellular Ca2+ dependence was non-linear when long depolarizing pulses (up to 500 ms) were applied; the amplitude of [Ca2+]i transient/Ca2+ entry (unit [Ca2+]i transient) increased with an increase in Ca2+ entry. 6. Increasing the duration of depolarization (-50 or -60 to 0 mV) from 20 to 500 ms enhanced asymptotically the integral of ICa (due to inactivation), and progressively the magnitude of [Ca2+]i transients, leading to the apparent non-linear dependence of unit [Ca2+]i transient on Ca2+ entry as well as on the duration of membrane depolarization. The peak time of [Ca2+]i transient was unchanged for pulse durations up to 300 ms, but prolonged with an increase in pulse duration to 500 ms. 7. Inhibitors of Ca2+ release from intracellular Ca2+ reservoirs, dantrolene (10 microM) and ryanodine (50 microM), blocked the [Ca2+]i transient to 56 and 30%, respectively, of the control. 8. The higher the basal [Ca2+]i level, the greater was the magnitude of the [Ca2+]i transients.(ABSTRACT TRUNCATED AT 400 WORDS)

Excitation-induced Ca2+ dynamics in sympathetic neurons measured with conventional epifluorescence and confocal UV laser-scanning microscopes.

A depolarization-induced rise in intracellular Ca2+ in cultured bullfrog sympathetic ganglion cells depended non-linearly on Ca2+ influx and membrane depolarization, spread inwardly faster than that without Ca(2+)-induced Ca2+ release (CICR), and was blocked by dantrolene and ryanodine. Ca2+ entry and a graded activation of CICR thus induce the "Ca2+ transient."

Intracellular calcium dynamics in response to action potentials in bullfrog sympathetic ganglion cells.

1. Dynamic changes in the intracellular free Ca2+ concentration ([Ca2+]i) following electrical membrane activity, were recorded from the neurone soma of the excised bullfrog sympathetic ganglion, using Fura-2 fluorescence and compared with the accompanying Ca(2+)-dependent electrical membrane responses. 2. The resting [Ca2+]i was about 100 nM, a value little changed by penetration with an intracellular electrode. 3. A net rise in fluorescence at a wavelength of 340 nm (Ca2+ transient) induced by a single action potential in Ringer solution rose almost in parallel with the initial decay phase of a slow Ca(2+)-dependent after-hyperpolarization; decayed in parallel with the late phase; and increased in amplitude and duration in the presence of tetraethylammonium (20 mM). 4. A Ca2+ transient induced by repetitive action potentials was increased asymptotically in amplitude and progressively in duration by increasing the number of spikes, and was slower in time course than the associated Ca(2+)-dependent K+ current. 5. Scanning a single horizontal line across the cytoplasm with an ultraviolet argon ion laser (351 nm) and recording Indo-1 fluorescence with a confocal microscope demonstrated an inward spread of a rise in [Ca2+]i following a tetanus. 6. Both single spike- and tetanus-induced Ca2+ transients were abolished in a Ca(2+)-free solution, while single or repetitive transient rises in [Ca2+]i induced by caffeine (5-10 mM) were generated under the same conditions. 7. Ryanodine (10-50 microM) did not affect tetanus-induced Ca2+ transients, whereas it blocked completely the caffeine-induced oscillation of [Ca2+]i. 8. Ca2+ transients induced by a tetanus in Ringer solution were independent of the interval from the preceding tetanus. The amplitude of Ca2+ transients induced by a tetanus in the presence of caffeine (5 mM) was equal to, or greater than, that generated in Ringer solution in any of the phases of [Ca2+]i oscillation. 9. It is suggested that under the physiological conditions here, the induction of action potentials does not cause the release of Ca2+ in the cells of the freshly excised bullfrog sympathetic ganglion, and that Ca(2+)-buffering systems contribute not only to lowering a transient rise in [Ca2+]i but also to sustaining an increased [Ca2+]i after a large Ca2+ load into the cell.

Basal Ca2+ and the oscillation of Ca2+ in caffeine-treated bullfrog sympathetic neurones.

1. Effects of caffeine on the intracellular free Ca2+ concentration ([Ca2+]i) in single bullfrog sympathetic neurones in excised tissue were studied by recording Fura-2 fluorescence excited at 340, 361 or 380 nm and taking their ratios (R340/380 or R361/380). 2. Caffeine (3-10 mM) produced oscillation of [Ca2+]i and an 'apparent' decrease in the basal level of [Ca2+]i during a period between phasic rises. The mechanism of the latter effect was analysed in relation to the mechanism of the former. 3. Caffeine (3-10 mM) increased Fura-2 fluorescence in a range of excitation wavelength from 330 to 390 nm. The ratios of fluorescences, R340/380 and R361/380, however, were not significantly affected by caffeine. These results suggest that the 'apparent' reduction in the basal [Ca2+]i seen as a decrease in R340/380 or R361/380 results from a true decrease in [Ca2+]i. 4. Caffeine-induced decrease in [Ca2+]i persisted for every period between phasic rises of [Ca2+]i during [Ca2+]i oscillation, and after the blockade of [Ca2+]i oscillation by ryanodine. The decrease in the latter condition lasted for more than 20 min. 5. The decrease in the basal [Ca2+]i depended on the external Ca2+ concentration and was not mimicked by the action of cyclic nucleotides. 6. Possible mechanisms underlying the decrease in the basal [Ca2+]i produced by caffeine (effects on Ca2+ transport at the cell or Ca(2+)-storing organelle membrane) and their significance in relation to the [Ca2+]i oscillation were discussed.

Intracellular Ca2+ dynamics in response to Ca2+ influx and Ca2+ release in autonomic neurones.

Spatial and temporal changes in the intracellular free Ca2+ concentration in response to Ca2+ influx at the cell membrane and to Ca2+ release from intracellular organelles were studied by recording fluorescence of Ca(2+)-sensitive probes, fura 2 or indo 1, with conventional epifluorescence or confocal laser-scanning microscopy combined with recordings of Ca(2+)-dependent membrane responses in bullfrog sympathetic ganglion cells. It was found that an increase in the intracellular Ca2+ induced by (an) action potential(s) in freshly ganglion cells bathed in Ringer's solution was solely a result of Ca2+ influx, while a rise in the intracellular Ca2+ by Ca2+ current in voltage-clamped cultured neurones was caused by not only Ca2+ influx but also Ca2+ release. This Ca2+ release was suggested to occur by a voltage-dependent (and graded) mode of activation of a Ca(2+)-induced Ca2+ release mechanism, explaining the lack of Ca2+ release by action potentials (because of their short-lasting depolarization) in freshly isolated neurons. In both cases, there was an inward spread of an increase in intracellular Ca2+. On the other hand, all or nothing activation of Ca(2+)-induced Ca2+ release occurred in the presence of caffeine, leading to the oscillation of Ca2+ in the cells. Characteristics of this mode of Ca2+ release and unique properties of drugs to block Ca2+ release were described. Finally, the physiological significance of different types of Ca2+ release was discussed.

Ultraviolet light activates blocking actions of dantrolene on intracellular Ca2+ release in bullfrog sympathetic neurones.

Effects of dantrolene, a blocker of intracellular Ca2+ release, on the oscillation of the intracellular Ca2+ ([Ca2+]i) induced by caffeine were studied in bullfrog sympathetic ganglion cells, using a Fura-2 fluorescence technique. Dantrolene blocked the Ca2+ oscillation only in the cell illuminated by ultraviolet light (335-385 nm). Likewise, the blocking effects on rhythmic Ca(2+)-dependent hyperpolarizations, representing Ca2+ oscillations via activation of Ca(2+)-dependent K+ channel, occurred only under the illumination with ultraviolet light (335-385 nm), but not with visible light (404-417 nm). This wavelength dependence differs from the absorbance spectrum of dantrolene. On the other hand, dantrolene preirradiated with ultraviolet light under dark condition or ultraviolet light itself did not affect the [Ca2+]i oscillation. The blocking action was not prevented by the pretreatment of the cells with reducing agents. These results indicate that illumination of the Ca2+ release channel or dantrolene itself with ultraviolet light (possibly the former) is necessary for the drug to exert its blocking effect. Furthermore, dantrolene was found to decrease Fura-2 fluorescence and to increase cell autofluorescence, leading sometimes to a false decrease in the basal [Ca2+]i.

Spatial and dynamic changes in intracellular Ca2+ measured by confocal laser-scanning microscopy in bullfrog sympathetic ganglion cells.

Confocal laser scanning microscopy (CLSM) was used to record spatial and dynamic changes in the intracellular Ca2+ [(Ca2+]i) of bullfrog sympathetic ganglion cells in excised tissue or in culture. A CLSM utilizing Ar ion laser (488 nm) and recording fluo-3 fluorescence yielded the sliced image of ganglion cells, while conventional epifluorescence microscopy provided the cell image of a convex structure. A high K+ (50 mM) solution, caffeine (3-10 mM) and electrical stimulation (10-20 Hz, 0.5-10 s) caused a homogeneous increase in fluo-3 fluorescence with or without regional differences, possibly due to intracellular organelles and other constituents. Scanning a single horizontal line across the cytoplasm with He-Cd laser (325 nm) and recording indo-1 fluorescence demonstrated that the rate of rise in [Ca2+]i following action potentials depends on the distance from the cell membrane and on the cytoplasmic constituents, showing an inward spread of 'Ca(2+)-wave' at variable speeds of 17-219 microns/s. These results suggest that heterogeneity of the cytoplasmic structures and constituents affects dynamic and spatial changes of [Ca2+]i in response to stimuli in neurones. Such heterogenic changes in [Ca2+]i would better be studied by CLSM.

Fast hyperpolarization following an excitatory postsynaptic potential in cat bladder parasympathetic neurons.

Intracellular recording techniques were used to study a fast hyperpolarizing potential following the fast excitatory postsynaptic potential evoked by an orthodromic nerve stimulation in cat bladder parasympathetic ganglion cells. In the 61 ganglion cells examined, two types of responses were recorded on stimulating the preganglionic nerve; one had only a fast excitatory postsynaptic potential (type SI, n = 20) and the other had a fast excitatory postsynaptic potential followed by a fast hyperpolarizing potential (type SII, n = 41). In type SII neurons, the half-maximum duration of the afterhyperpolarizing potential following an orthodromic spike was longer than that of a direct spike produced by injecting a depolarizing current pulse through the recording electrode; the half-maximum durations for afterhyperpolarizing potentials following orthodromic and direct action potentials were comparable in type SI cells. Blocking the initiation of an orthodromic spike by hyperpolarizing the membrane in type SII cells revealed a fast excitatory postsynaptic potential followed by a fast hyperpolarizing potential which was similar to that observed at the resting potential. The fast hyperpolarizing potential had a duration comparable to that of an afterhyperpolarizing potential following an orthodromic action potential. The fast excitatory postsynaptic potential-fast hyperpolarizing potential sequence was blocked completely and reversibly by nicotinic receptor antagonists (hexamethonium and D-tubocurarine). Atropine, alpha-2 noradrenergic (yohimbine and phentolamine), and purinergic (caffeine) antagonists had no effect on the fast hyperpolarizing potential. In cells which show type SII responses, spontaneous excitatory postsynaptic potentials were not followed by a hyperpolarization. Depolarizing the membrane (by passing a cathodal current through the recording electrode) to an amplitude comparable to that of a fast excitatory postsynaptic potential also did not elicit a membrane hyperpolarization in type SII cells. In some cells, stimulating one preganglionic nerve trunk elicited a fast hyperpolarizing potential, but activating another nerve trunk innervating the same ganglion cell did not. There was no correlation between the variations in the amplitudes of the fast excitatory postsynaptic potential and the fast hyperpolarizing potential in type SII cells, but increasing the stimulus intensity applied to the presynaptic nerve fiber potentiated the amplitude of the fast excitatory postsynaptic potential and the fast hyperpolarizing potential. The fast hyperpolarizing potential was not associated with appreciable changes in input resistance.(ABSTRACT TRUNCATED AT 400 WORDS)

Measurement of intracellular Ca2+ in the bullfrog sympathetic ganglion cells using fura-2 fluorescence.

The intracellular free Ca2+ concentration ([Ca2+]i) of the bullfrog sympathetic ganglion cell was measured with fura-2 fluorescence under various conditions, and compared with changes in membrane potential recorded with an intracellular electrode. The [Ca2+]i was 109 nM on average under the resting condition and increased by raising the extracellular K+, stimulating repetitively the pre- or post-ganglionic nerve, or by applying acetylcholine or muscarine. Since all these procedures depolarized the cell membrane, most of the rise in [Ca2+]i could be the result of opening of voltage-dependent Ca2+ channels. However, Ca2+ entries through nicotinic acetylcholine receptor channels and the channel activated by the muscarinic acetylcholine receptor were also indicated by considering the threshold for the opening of voltage-dependent Ca2+ channels (for both entries) or a limited number of the cells showing the latter response.

Role of ion conductance changes and of the sodium-pump in adrenaline-induced hyperpolarization of rat diaphragm muscle fibres.

The ionic mechanism of membrane hyperpolarization induced by adrenaline in rat diaphragm muscle fibres was studied. Removal of the extracellular K+ ([K+]o) from Krebs-Ringer solution initially increased the resting membrane potential and then caused an increase in the intracellular Na+ activity ([Na+]i) and a decrease in the intracellular K+ activity ([K+]i). All the changes were maintained for more than 3 h. Application of ouabain (0.1 mM) or lowering the temperature rapidly reduced the resting potential by about 10 mV in the K+-free solution. It then produced further progressive decreases in resting potential and in [K+]i and a progressive increase in [Na+]i. These observations indicate that an electrogenic Na-pump operates in the K+-free solution. Removal of most of the Cl- in the K+-free solution did not affect the resting potential or the magnitude of the initial decrease produced by ouabain, despite an increased input resistance; this result implies a passive distribution of Cl-. Adrenaline (30-60 microM) either added to the bathing solution or applied to the membrane by ionophoresis produced a hyperpolarization (3-10 mV: adrenaline hyperpolarization), the amplitude of which was decreased with a rise in [K+]o and increased with a reduction in [K+]o, but unaffected by the removal of Cl-. Adrenaline produced an increase in input resistance, the relative magnitude (17-18%) of which was constant whether external K+ or Cl- was removed. In contrast, a conditioning membrane hyperpolarization hardly affected the resistance. Ouabain (0.1 mM) or low temperature (8-10 degrees C) abolished both the hyperpolarization and the increased input resistance induced by adrenaline. The [K+]i, [Na+]i and the peak of the action potential remained unchanged after a 20 min exposure to adrenaline (30 microM). The hyperpolarization induced by the replacement of all Na+ with Tris (Tris-hyperpolarization) in the K+-free solution was depressed by 39% during the early period (4-31 min) of exposure to adrenaline (30 microM), while it was enhanced by 26% during the later period (80-130 min). The initial depression suggested a decrease in the ratio of the membrane permeability for Na+ (PNa) to that for K+ (PK). These results suggest that the adrenaline hyperpolarization is generated largely by a decrease in PNa/PK, which is associated with the activity of the Na-pump.

Characterization of gamma-aminobutyric acid responses with sulfate loading in cat bladder neurons.

In some cells in cat bladder ganglia gamma-aminobutyric acid (GABA) applied iontophoretically produced a hyperpolarizing response accompanied by an increase in input conductance when recorded with potassium sulfate-filled microelectrodes. This GABA-induced hyperpolarization was blocked by bicuculline and was converted to a depolarizing GABA response when extracellular chloride concentration was low suggesting that the hyperpolarizing GABA response was mediated by the opening of chloride channels. In other cells, continuous passage of a small negative current converted a depolarizing GABA response to a hyperpolarizing response with time. This effect was accompanied by a negative shift of the reversal potential. These data indicated that injection of impermeable sulfate ions decreased the intracellular chloride concentration.

Calcitonin and calcitonin gene-related peptide enhance calcium-dependent potentials.

Recent data suggests that calcitonin (CT) and/or calcitonin gene-related peptide (CGRP) may be potential transmitters or modulators in the nervous system. The present study analyzed the effect of CT and CGRP on the neuronal membranes of cat parasympathetic ganglia of the urinary bladder. The related peptides prolonged the duration of the afterhyperpolarization of the action potential but had no effect on resting potential or input resistance. CT and CGRP enhanced the duration of a calcium spike recorded in the presence of agents blocking Na and K channels while under similar conditions forskolin, an activator of adenylate cyclase, did not affect the calcium spike. These data suggest that the neural mechanism of action of CT and CGRP is to prolong a calcium conductance and that these effects are not mediated through cyclic AMP.

Effects of Na+ gradient on the intracellular Ca2+ oscillation in the sympathetic ganglion cell: Na-Ca exchange in the neurone cell soma?

The effects of the removal of extracellular Na+ and the increase in the intracellular Na+ on the slow rhythmic membrane hyperpolarizations (rmhs) were studied in bullfrog sympathetic ganglion cells. The interval of rmhs, reflecting a relative change in the basal intracellular Ca2+, was lengthened by either treatment, disfavouring the possible involvement of the Na-Ca exchange mechanism in the Ca2+-buffering in the ganglion cell soma.

(+)-Tubocurarine blocks the Ca2+-dependent K+-channel of the bullfrog sympathetic ganglion cell.

(+)-Tubocurarine [+)-Tc: 10-100 microM) reduced the duration of the afterhyperpolarization, which was induced by the activation of Ca2+-dependent K+-conductance (GK,Ca) following an action potential in the bullfrog sympathetic ganglion cell, but did not affect the maximum rates of rise and fall of Na+- and Ca2+-dependent action potentials. The amplitudes of slow rhythmic membrane hyperpolarizations produced by rhythmic rises in the GK,Ca were also decreased by (+)-Tc without a change in their intervals. Thus, (+)-Tc appears to block the Ca2+-dependent K+-channel of the bullfrog sympathetic ganglion cell.