Knockout of the BK β2 subunit abolishes inactivation of BK currents in mouse adrenal chromaffin cells and results in slow-wave burst activity

Rat and mouse adrenal medullary chromaffin cells (CCs) express an inactivating BK current. This inactivation is thought to arise from the assembly of up to four β2 auxiliary subunits (encoded by the kcnmb2 gene) with a tetramer of pore-forming Slo1 α subunits. Although the physiological consequences of inactivation remain unclear, differences in depolarization-evoked firing among CCs have been proposed to arise from the ability of β2 subunits to shift the range of BK channel activation. To investigate the role of BK channels containing β2 subunits, we generated mice in which the gene encoding β2 was deleted (β2 knockout [KO]). Comparison of proteins from wild-type (WT) and β2 KO mice allowed unambiguous demonstration of the presence of β2 subunit in various tissues and its coassembly with the Slo1 α subunit. We compared current properties and cell firing properties of WT and β2 KO CCs in slices and found that β2 KO abolished inactivation, slowed action potential (AP) repolarization, and, during constant current injection, decreased AP firing. These results support the idea that the β2-mediated shift of the BK channel activation range affects repetitive firing and AP properties. Unexpectedly, CCs from β2 KO mice show an increased tendency toward spontaneous burst firing, suggesting that the particular properties of BK channels in the absence of β2 subunits may predispose to burst firing

Nicotonic and muscarinic components in acetylcholine stimulation of porcine adrenal medullary cells

Adrenal medullary chromaffin cells secrete catecholamines (CA) in response to cholinergic receptor activation by acetylcholine (ACh) released from splacnic nerve terminals. In cultured bovine chromaffin cells nicotinic receptors play a preponderant (> 90%) role in the control of CA release. By contrast, we found and report here that up to 40% of the ACh-evoked CA secretion from cultured porcine chromaffin cells can be associated with muscarinic receptor activation. The following results support our belief that in porcine adrenal medullary cells ACh (100 μM) evoked CA secretion is mediated by both nicotinic and muscarinic cholinergic receptors. 1) Hexamethonium (100 μM), a nicotinic receptor antagonist, inhibited ACh-induced CA secretion to ca. 40% of the control release and atropine (1 μM), a muscarinic receptor antagonist, inhibited to ca. 60% of the control value. 2) We also found that ACh(100 μM) evoked intracellular Ca2+ concentration ([Ca2+](i)) rise was inhibited by these recept

Deprivation of Na +, Ca 2+ and Mg 2+ from the extracellular solution increases cytosolic Ca 2+ and stimulates catecholamine secretion from cultured bovine adrenal chromaffin cells

We report here that exposing cultured chromaffin cells to a low ionic strength medium (with sucrose in place of NaCl to maintain osmolarity) can induce a marked elevation in cytosolic Ca 2+ concentration ([Ca 2+](i)) and catecholamine (CA) release. To determine the underlying mechanism, we first studied the effects of low [Na +](o) on single cell [Ca 2+](i) (using fluo-3 as Ca 2+ indicator) and CA release from many cells. In a Mg 2+ and Ca 2+-deficient medium, lowering the external concentration of Na 2+ ([Na +](o)) evoked CA secretion preceded by a transitory [Ca 2+](i) rise, the amplitude of which was inversely related to [Na +](o). By contrast, in the presence of either [Ca 2+](o) (2 mM) and [Mg 2+](o) (1.4 mM) or [Mg 2+](o) alone (3.4 mM), lowering the ionic strength was without effect. Furthermore, in a physiologic [Na +](o), [Ca 2+](o) and [Mg 2+](o) medium, two or three consecutive applications of the cholinergic agonist oxotremorine-M (oxo-M) consistently evoked a substantial

Metabolic correlates of fatigue and of recovery from fatigue in single frog muscle fibers

Fatigue and recovery from fatigue were related to metabolism in single fibers of the frog semitendinosus muscle. The fibers were held at a sarcomere length of 2.3 µm in oxygenated Ringer solution at 15°C and were stimulated for up to 150 s by a schedule of 10-s, 20-Hz tetanic trains that were interrupted by 1-s rest periods, after which they were rapidly frozen for biochemical analysis. Two kinds of fatigue were produced in relation to stimulus duration. A rapidly reversed fatigue occurred with stimulation for under 40 s and was evidenced by a decline in tetanic tension that could be overcome by 1 s of rest. A prolonged fatigue was caused by stimulation for 100-150 s. It was evidenced during stimulation by a fall in tetanic tension that could not be overcome by 1 s of rest, and after stimulation by a reduction, lasting for up to 82 min, in the peak tension of a 200-ms test tetanus. Fiber phosphocreatine (PCr) fell logarithmically in relation to stimulus duration, from a mean of 121 ±