Whole cell patch-clamp recordings of rat midbrain dopaminergic neurons isolate a sulphonylurea- and ATP-sensitive component of the potassium currents activated by hypoxia.. E. Guatteo, M. Federici, A. Siniscalchi, T. Kn”pfel, N. B. Mercuri and G. Bernardi. IRCCS Santa Lucia and Clinica Neurologica, Universita' di Tor Vergata, Roma, Italy.
APStracts 4:349N, 1997.
ABSTRACT
The effects of brief (2 - 4 min) hypoxia on presumed dopaminergic "principal" neurons of the rat ventral mesencephalon were investigated using either intracellular or whole-cell patch-clamp recordings in in vitro conditions. Under single-electrode voltage-clamp, with sharp microelectrode (V_h_ -60 mV), a brief hypoxia caused an outward current (hypo-_out_) of 110.2 ñ 15.2 pA (n = 18) which was followed by a post-hypoxic outward current (post- hypo-_ out_) of 149.6 ñ 10.6 pA (n = 18). While the hypo-_out_ reversed at -83.7 ñ 3.8 mV (n = 18), the post- hypo-_out_ did not reverse. The K_+_ATP_-blocking sulphonylureas tolbutamide (100 æM) and glibenclamide (30 æM), significantly reduced the peak of the hypo-_out_ by 47.6 ñ 7.7 % (n = 16) and 54.18 ñ 7.5 % (n = 3), respectively. In contrast, they did not affect the post-hypo-_out_. Extracellular barium (300 æM - 1 mM) almost abolished the hypo-_out_, leaving the post-hypo-_ out_ unchanged. The large K_+_ channel blocker charybdotoxin (10 - 50 nM), depressed the hypo-_out_ after tolbutamide treatment. In order to investigate whether cytosolic factors might control the development of the hypo-_out_, we dialyzed the principal neurons by patch- clamp recordings (V_h_ -60 mV). Under whole-cell recordings hypoxia evoked an hypo-_out_ of 70.2 ñ 14.5 pA that reversed polarity at -87.9 ñ 5.1 mV (n = 8). A small post-hypoxic response was detected upon re-oxygenation in a few neurons (4 out of 14). Three different sulphonylureas, tolbutamide (100 æM), glibenclamide (10 - 30 æM) and glipizide (100 nM) completely blocked the hypo- _out_ in patch-clamped neurons. The hypo-_out_ was also abolished by extracellular BaCl2 (300 æM). When the content of ATP in the dialyzate was raised from 2 mM to 10 mM no outward current/hyperpolarization was evoked by hypoxia. These data suggest that the hypo-_out_, in principal neurons, is a complex response sustained by at least two barium-sensitive components: 1) an ATP- dependent, sulphonylurea-sensitive K_+_ conductance which could be isolated by the patch-clamp techniques. 2) a K_+_ conductance remaining after tolbutamide in intracellularly recorded neurons, which is sensitive to charybdotoxin and dependent on dialyzable cytosolic factors. 

Received 9 September 1997; accepted in final form 3 December 1997.
APS Manuscript Number J739-7.
Article publication pending J. Neurophysiol.
ISSN 1080-4757 Copyright 1997 The American Physiological Society.
Published in APStracts on 12 December 1997