EICOSANOIDS MODULATE APICAL CA-2+ DEPENDENT K+ CHANNELS IN CULTURED RABBIT PRINCIPAL CELLS

Patch clamp technology was utilized to study the effects of apical phospholipase A2 (PLA2) metabolites on "maxi K" channels in the principal cell apical membrane of rabbit cortical collecting tubule (CCT) primary cultures (B. N. Ling, C. F. Hinton, and D. C. Eaton. Kidney Int. 40: 441-452, 1991). At resting membrane potential, this channel is quiescent in the cell-attached configuration. Apical application of the PLA2 agonist melittin (1-mu-g/ml) for 10 min increased single-channel open probability (P(o)) from 0.0004 +/- 0.0010 to 0.11 +/- 0.05. Similarly, apical exposure to 50-mu-M arachidonic acid (AA) or 0.5-mu-M prostaglandin (PG) E2, but not 0.5-mu-M PGF2-alpha, also increased channel activity. Conversely, 10-mu-M of the PLA2 antagonist quinacrine applied apically decreased P(o). Removal of apical bath Ca2+ did not prevent melittin-, AA-, or PGE2-induced channel activation. We then examined the role of maxi K channels and eicosanoids in principal cell volume regulation. Within seconds of reducing basolateral bath tonicity (285 to 214 mosmol/kgH2O), Np(o) (i.e., no. of channels x P(o)) initially increased almost-equal-to 200%, followed by a delayed but prolonged activation phase that was attenuated by removal of apical bath Ca2+. Pretreatment with 10-mu-M quinacrine, 100-mu-M indomethacin (cyclooxygenase inhibitor), or 0.25-mu-M thapsigargin (to deplete intracellular Ca2+ stores) abolished the initial phase of swelling-induced channel activation. We concluded the following: 1) AA, cleaved from apical membrane phospholipids by PLA2, opens maxi K channels in rabbit CCT principal cells; 2) channel activation by AA and PGE2 occurs independent of extracellular Ca2+, whereas full activation by principal cell swelling requires luminal Ca2+ influx; and 3) modulation of distal nephron K+ efflux by apical membrane phospholipids may play an important physiological role in principal cell volume homeostasis.