Low-threshold Ca Currents Research Articles

Graded Inhibitory Synaptic Transmission Between Leech Interneurons: Assessing the Roles of Two Kinetically Distinct Low-Threshold Ca Currents

In leeches, two pairs of reciprocally inhibitory heart interneurons that form the core oscillators of the pattern-generating network for heartbeat possess both high- and low-threshold (HVA and LVA) Ca channels. LVA Ca current has two kinetically distinct components (one rapidly activating/inactivating, ICaF, and another slowly activating/inactivating, ICaS) that mediate graded transmission, generate plateau potentials driving burst formation, and modulate spike-mediated transmission between heart interneurons. Here we used different stimulating protocols and inorganic Ca channel blockers to separate the effects of ICaF and ICaS on graded synaptic transmission and determine their interaction and relative efficacy. Ca2+ entering by ICaF channels is more efficacious in mediating release than that entering by ICaS channels. The rate of Ca2+ entry by LVA Ca channels appears to be as critical as the amount of delivered Ca2+ for synaptic transmission. LVA Ca currents and associated graded transmission were selectively blocked by 1 mM Ni2+, leaving spike-mediated transmission unaffected. Nevertheless, 1 mM Ni2+ affected homosynaptic enhancement of spike-mediated transmission that depends on background Ca2+ provided by LVA Ca channels. Ca2+ provided by both ICaF and ICaS depletes a common pool of readily releasable synaptic vesicles. The balance between availability of vesicles and Ca2+ concentration and its time course determine the strength of inhibitory transmission between heart interneurons. We argue that Ca2+ from multichannel domains arising from ICaF channels, clustered near but not directly associated with the release trigger, and Ca2+ radially diffusing from generally distributed ICaS channels interact at common release sites to mediate graded transmission.

Intracellular Ca2+Dynamics During Spontaneous and Evoked Activity of Leech Heart Interneurons: Low-Threshold Ca Currents and Graded Synaptic Transmission

In oscillatory neuronal networks that pace rhythmic behavior, Ca(2+) entry through voltage-gated Ca channels often supports bursting activity and mediates graded transmitter release. We monitored simultaneously membrane potential and/or ionic currents and changes of Ca fluorescence (using the fluorescence indicator Ca Orange) in spontaneously active and experimentally manipulated oscillator heart interneurons in the leech. We show that changes in Ca fluorescence in these interneurons during spontaneous bursting and evoked activity reflect the slow wave of that activity and that these changes in Ca fluorescence are mediated by Ca(2+) entry primarily through low-threshold Ca channels. Spatial and temporal maps of changes in Ca fluorescence indicate that these channels are widely distributed over the neuritic tree of these neurons. We establish a correlation between the amount of transmitter released, as estimated by the integral of the postsynaptic current, and the change in Ca fluorescence. In experiments in which we were able to record presynaptic low-threshold Ca currents, associated IPSCs, and presynaptic changes in Ca fluorescence from fine neuritic branches of heart interneurons near their region of synaptic contact with their contralateral partner, there was a close association between the rise in Ca fluorescence and the rise of the postsynaptic conductance. The changes in Ca fluorescence that we record at the end of fine neuritic branches appear to reflect changes in [Ca(2+)](i) that mediate graded synaptic release in leech heart interneurons. These results indicate that widely distributed low-threshold Ca currents play an important role in generating rhythmic activity and in mediating graded transmitter release.

Benzolamide inhibits low-threshold calcium currents in hippocampal pyramidal neurons.

1. Benzolamide is a poorly permeant sulfonamide inhibitor of the enzyme carbonic anhydrase. We studied the effect of benzolamide on low-threshold (LT) Ca currents in neonatal hippocampal CAl neurons. 2. In hippocampal slices, benzolamide (2-10 microM) inhibited the LT current 30-75% in voltage-clamped CAl pyramidal cells (n = 6). In slices bathed in N-2-hydroxypiperazine-N'-2-ethane-sulfonic acid (HEPES)-buffered Ringer, benzolamide also reduced the LT current, indicating that the action of the drug was not bicarbonate dependent. 3. Benzolamide inhibited LT Ca currents 20-75% in acutely dissociated CAl neurons in HEPES (n = 18): inhibition was 36 +/- 8% (mean +/- SE; n = 7) and 50 +/- 8% (n = 7) at 10 and 50 microM benzolamide, respectively. By contrast, high-threshold calcium currents recorded in CAl pyramidal cells (n = 18) and dorsal root ganglion neurons (n = 4) were virtually unaffected by benzolamide. 4. These results indicate that benzolamide inhibits LT Ca channels in central neurons and suggest caution in the use of this agent to inhibit extracellular carbonic anhydrase in excitable tissues.

Low-threshold calcium current in dendrites of the adult rat hippocampus

The patch-clamp technique was employed in hippocampal slices to examine the characteristics of a low-threshold Ca current in adult CA1 pyramidal neurons. We found that adult CA1 pyramidal neurons possess a distinct transient low-threshold Ca current, located predominantly in the distal dendrites. Surgical cuts that separated the dendrites from the soma and left a dendritic length of 150 μm, completely abolished the low-threshold Ca current while a transient K current persisted even in cells with dendrites as short as 50 μm. The transient low-threshold Ca current in dendrites may represent a voltage-dependent Ca entry pathway which contributes to the regulation of cellular excitability, plasticity and pathology.

Modulation by neuromediators of the low threshold Ca current, I CaT in single bullfrog atrial cells

Modulation by neuromediators of the low threshold Ca current, I CaT in single bullfrog atrial cells

Relationships between capsaicin sensitivity of mammalian sensory neurons, cell size and type of voltage gated Ca-currents

(1) The relationship between capsaicin sensitivity, cell size and the expression of voltage-dependent high-threshold (L- and N-type) and low-threshold (T-type) Ca-currents were investigated in cultured adult rat dorsal root ganglion neurons using the whole-cell patch-clamp technique. (2) The capsaicin (10 μM) evoked inward current was used to identify the capsaicin-sensitive population. To identify the voltage-dependent Ca-currents, the activation threshold and the inactivation kinetic were used. (3) In cultured dorsal root ganglion cells the capsaicin sensitivity is not restricted to the small cell population. About 28% of cells with large cross sectional areas (⩾1001 μm 2) responded with about the same amplitude per μm 2 cross sectional area as small cells. (4) The amplitude of the capsaicin evoked inward current depended on the length of time the cells were maintained in culture. (5) All cells exhibited the high-threshold Ca-currents. About 45% of the c capsaicin-insensitive cells also had low-threshold Ca-currents. None of the capsaicin-sensitive cells had the low-threshold type. (6) Most of the cells which expressed the low-threshold type were in the upper size range.

Regulation of ionic currents in pheochromocytoma cells by nerve growth factor and dexamethasone

Growth factors and hormones induce differentiation of clonal pheochromocytoma (PC12) cells, which are derived from rat adrenal medulla chromaffin cells. On application of nerve growth factor (NGF), PC12 cells extend neurites and express properties characteristic of autonomic ganglion cells. In contrast, incubation of PC12 cells with a corticosteroid, dexamethasone (DEX), does not induce neurite formation but causes an increase in tyrosine hydroxylase activity, suggesting that the cells become chromaffin cell-like. The ability of NGF and DEX to regulate ionic currents has been less well studied. Therefore, we examined how long-term NGF and DEX treatments affected voltage-dependent Na, Ca, and K currents in PC12 cells. Voltage-dependent Na currents were observed only in a small fraction of the PC12 cells in the absence of NGF or DEX. Virtually all NGF-treated cells expressed Na currents within 7 d. DEX increased the number of cells expressing voltage-dependent Na current slowly over 3 weeks, but, unlike NGF, DEX did not change Na current density. Both NGF and DEX also affected the expression of voltage-dependent Ca currents. Most of the untreated cells had only sustained, high-threshold voltage-dependent Ca currents. Chronic application of NGF or DEX increased the fractions of the cells that showed transient, low-threshold T-type Ca currents in addition to the high-threshold currents. The T-type Ca current density, however, increased significantly only in NGF-treated cells. Neither DEX nor NGF affected the voltage-dependent K currents. These results suggest that the expression of voltage-dependent Na and Ca currents are differentially regulated by NGF and DEX. The distinction between treated and untreated cells decreased after 3 weeks in culture as older untreated cells showed increases in the fraction of cells expressing both Na and low-threshold Ca currents. A PC12 subline selected for adherence to uncoated plastic also showed increased fraction of cells expressing these currents, suggesting that interactions with substrate may also influence ionic current expression.

The low-threshold Ca current in isolated amygdaloid neurons in the rat

Two types of voltage-dependent Ca currents were recorded from isolated rat amygdaloid neurons under single-electrode voltage-clamp. A low-threshold Ca current was elicited at −60 mV or more positive potentials and inactivated rapidly. At −30 mV or more positive potentials, a high-threshold Ca current was also activated. In the steady-state inactivation curve of the low-threshold Ca current, the half-inhibition value (h 0.5) was −71 mV. The low-threshold Ca current was inhibited by organic and inorganic Ca blockers in a dose-dependent manner. The inhibitory effect of these Ca blockers was completely reversible while that of flunarizine was partly so. It is concluded that the membrane and pharmacological properties of the low-threshold Ca channel in the rat amygdaloid neurons are quite similar to those in the hypothalamic neurons.

Low-threshold calcium electrogenesis in neocortical neurons

In slices of parietal neocortex, evidence was obtained for the existence of a low-threshold Ca 2+ conductance in most neurons. This conductance became apparent when resting membrane potential was held below −60 mV by continuously injected, depolarizing current. Under these conditions, brief hyperpolarizing pulses were followed by generation of tetrodotoxin (TTX)-resistent, Mn 2+-sensitive, low-threshold spikes. The results suggest that in neocortex, as in many subcortical structures, low-threshold Ca current may be responsible for burst generation in some neurons.

Effect of menthol on two types of Ca currents in cultured sensory neurons of vertebrates

The effect of menthol on voltage-dependent Ca currents was investigated in cultured dorsal root ganglion cells from chick and rat embryos. Bath application of menthol (0.1-1 mM) had different effects on the various Ca currents present in these neurons. Below -20 mV, the low threshold Ca currents were reduced in amplitude in a dose-dependent manner by menthol with little changes of their activation kinetics. In contrast to this, the time course of inactivation of the high-threshold Ca currents, activated above -20 mV from a holding potential of -80 mV, was drastically accelerated by external menthol. The action of menthol was unchanged with more positive holding potentials (-50 mV). Thus, a proposed third type of Ca current with transient activation and complete deactivation below -50 mV was either not present or not affected by menthol. Menthol exerted its action only when applied from the outside. Its effect was completely reversible within 15-20 min of wash-out. Our findings are consistent with the idea that menthol acts on two types of Ca channels coexisting on the membrane of cultured sensory neurons. Menthol blocks currents through the low voltage-activated Ca channel, and facilitates inactivation gating of the classical high voltage-activated Ca channel.