Hyperpolarization-activated inward potassium and calcium-sensitive chloride currents in beating pacemaker insect neurosecretory cells (dorsal unpaired median neurons)

Hyperpolarization-activated inward currents were studied in single adult co ckroach Periplaneta americana pacemaker neurosecretory cells, identified as dorsal unpaired median neurons using the whole-cell patch-clamp technique. Under current clamp, injection of negative current produced st hyperpolari zation of the cell membrane with a sag in the membrane potential toward the resting value. Under voltage clamp, the whole-cell current-voltage relatio nship exhibited an unexpected biphasic aspect. The global hyperpolarization -activated inward current could be dissociated by means of 4-acetamido-4'-i sothiocyanostilbene-2,2'-disulfonic acid and tetraethylammonium chloride se nsitivity, ionic selectivity, voltage dependence and activation threshold a s inward potassium and calcium-sensitive chloride currents. The inward pota ssium current was activated around -80 mV. The reversal potential followed the potassium equilibrium potential when the extracellular potassium concen tration was raised. This current was not dependent on the external sodium c oncentration and was sensitive to 10 mM tetraethylammonium chloride or 5 mM barium chloride. The hyperpolarization-activated inward calcium-sensitive chloride current was activated in a range of potential 20 mV more positive than the potassium current. The estimated reversal potential (-71 mV) was v ery close to the equilibrium potential for chloride ions (-73 mV). Intracel lularly applied 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid, 4 ,4'-diisothiocyanostilbene-2,2'-disulfonic acid and external application of 1 mM zinc chloride, calcium-free saline or high concentrations of intracel lular 1,2-bis-(2-aminophenoxy) ethane-N,N,N',N'-tetra-acetate blocked the i nward chloride current. Current-clamp experiments indicated that the inward potassium current accounted for inward rectification of dorsal unpaired me dian neurons. Our findings report, for the first time in pacemaker neurosecretory cells, the co-existence of two distinct hyperpolarization-activated inward current s which have specialized function in pacemaker activity. (C) 1999 IBRO. Pub lished by Elsevier Science Ltd.