Apamin-sensitive Currents Research Articles

A novel calcium-activated apamin-insensitive potassium current in pituitary gonadotrophs.

In cultured rat pituitary gonadotrophs, GnRH-induced oscillations in cytosolic calcium concentration ([Ca2+]i) are associated with periodic membrane hyperpolarization. The hyperpolarizing waves are secondary to the activation of apamin-sensitive Ca2+-activated K+ channels that account for a single class of 125I-apamin binding sites present in these cells. In a substantial fraction of gonadotrophs, however, we observed a Ca2+-controlled oscillatory current that was resistant to apamin, even at concentrations five orders of magnitude higher than the dissociation constant (Kd) observed in the binding experiments. With the K+ in the pipette, the apamin-resistant current showed a reversal potential of -42 mV, nearly 40 mV more positive than that of the apamin-sensitive current. With Cs+ in place of K+ in the pipette solution, both the size of the apamin-insensitive current and its reversal potential remained unchanged. Ion substitution studies further revealed that the reversal potential was independent of Cl-. In contrast, an 11 mV hyperpolarizing shift in the reversal potential occurred when extracellular Na+ was reduced to 80 mM. In cells expressing apamin-resistant conductances, addition of apamin evoked a marked increase in the duration of the action potentials and reduction in the frequency of spontaneous spiking. In the presence of GnRH, gonadotrophs exhibit the typical burst pattern of electrical activity. Further exposure of the cells to apamin depolarized the membrane from a silent phase bursting level of about -80 mV to a new level of about -40 mV. These observations indicate that, in addition to apamin-sensitive current, a subpopulation of pituitary gonadotrophs also expresses a cationic component of the Ca2+-activated membrane conductance that has the potential to remodulate spontaneous and agonist-induced electrical activity.

Components of depolarization-induced transmembrane ion current in isolated smooth muscle cells of the guinea pigtaenia coli

Transmembrane ion currents in isolated single smooth muscle cells (SMC) from the guinea pigtaenia coli were investigated using a whole-cell mode of the patch-clamp technique. Currents induced by depolarizing shifts in the membrane potential from its holding level of −60 mV contained an initial inward phase (Ca2+ current), which in 30–40 msec was followed by an outward phase. It was shown that outward current was carried by K ions and consisted at least of three components: one Ca2+-independent K+ current of delayed rectifier (KV) and two Ca2+-dependent K+ currents. The latter can be further divided into the apamin-sensitive (SK) and charybdotoxin-sensitive (BK) currents. It was found that relative contributions of these three components in total outward current at 0 mV were 35–45%, 5–15%, and 45–55% for KV, SK, and BK currents, respectively. A potential-dependent current carried by Ci ions was also found. This Cl− current had inward direction within the range of potentials below the chloride equilibrium potential (E Cl) and outward direction above theE Cl. The magnitude of Cl− current was significantly lower than the magnitude of total K+ current.

Two distinct calcium‐activated potassium currents in a rat anterior pituitary cell line.

1. The single 'giga-seal' patch-electrode technique (Hamill, Marty, Neher, Sakmann & Sigworth, 1981) was used to record whole-cell currents in the GH3 rat anterior pituitary cell line. 2. GH3 cells have a rapidly inactivating, voltage-dependent K+ current that is selectively inhibited by 4-aminopyridine (4-AP) but not by tetraethylammonium chloride (TEA). 3. The majority of the Ca2+-activated K+ current in these cells is blocked by TEA with an inhibitory concentration that is half-maximal at 1 mM. An additional Ca2+-activated K+ current is also present that is relatively resistant to TEA and is blocked by the polypeptide apamin. The apamin-sensitive component represents less than 18% of the total Ca2+-activated K+ current at 0 mV. 4. The time course of the slowly declining components of the Ca2+-activated K+ tail currents measured at the -50 mV holding potential was usually biexponential with time constants of 0.21 +/- 0.02 and 1.75 +/- 0.23 s (mean +/- S.E. of mean, n = 14). Both of the two slowly decaying components contribute to the TEA- and apamin-sensitive currents. 5. It is concluded that GH3 cells have at least two pharmacologically distinct Ca2+-activated K+ currents and a 4-AP-sensitive voltage-dependent K+ current.

The apamin-sensitive potassium current in frog skeletal muscle: its dependence on the extracellular calcium and sensitivity to calcium channel blockers

Slow outward potassium currents were recorded in isolated frog skeletal muscle fibres using the double mannitol-gap voltage-clamp technique. Detubulated fibres failed to generate a slow outward current, and apamin had no effect on the remaining current. The maximum blocking effect of organic and inorganic Ca2+-channel blockers on the slow outward channels of intact fibres was larger than that of apamin. Apamin failed to induce an additional block when applied after Ca2+-channel blockers. In a low-Ca2+ solution (OCa, EGTA 1 mM) the slow outward current was slightly increased and the blocking effect of apamin was enhanced. A Ca2+-rich solution (Ca2+ X 10) increased the slow outward current and the blocking effect of apamin was drastically reduced. It is concluded that the apamin-sensitive current which is a component of the slow outward K+ current is located in the tubular membrane. Its activation seems barely dependent on the Ca2+ influx via the slow inward Ca2+ current. Apamin-receptor binding appears to be dependent on the extracellular Ca2+ concentration. Blockade of slow outward current by Ca2+-channel blockers is likely to be the result of a direct action on the slow K+ permeability rather than a consequence of Ca2+ channel inhibition.