ACTIVATION OF SMOOTH MUSCLE BK CHANNELS BY HYDROCHLOROTIAZIDE REQUIRES CELL INTEGRITY AND PRESENCE OF BK BETA-1 SUBUNITS.

ORSI, F; FLOREZ CASTRO, A; ASUAJE, A; MILESI, V; MARTÍN, P

Buenos Aires

Congreso; Reunión Conjunta de Sociedades de Biociencias; 2017

Sociedad Argentina de Investigación Clínica (SAIC)

Hydrochlorothiazide (HCTZ) is used to treat hypertension, their efficacyis linked to a chronic vasodilatory effect. Previous studies suggest thatactivation of the large conductance voltage- and Ca2+-dependent K+(BK) channel is responsible for HCTZ-induced vasodilatory effect. However, directelectrophysiological evidence supporting this claim is lacking.  BK channels can be associated with accessoryβ-subunits, which confer specific biophysical and pharmacologicalcharacteristics. The β1-subunit is mainly expressed in smooth musclecells (SMCs). Methods: We evaluatedthe effect of HCTZ on BK channel activity using patch-clamp technique on SMCsfrom human umbilical artery (HUASMCs) and in HEK293T cells expressing the BKchannel. Paired t-tests were used to compare two groups. Results: Using the whole-cell configuration (WCC) in HEK cells expressingthe BK channel with β1-subunit we observed that HCTZ raised thecurrent amplitude with an EC50 of 28.4 µM (pD2=4.5 ± 0.2, n:8).However, 30 µM HCTZ did not change the channel activity when it was evaluatedin the same cells in the inside-out configuration (IOC), where cell integrityis lost (%current increase (+90 mV): 12.8 ± 38.1, P>0.05; n:4), suggestingan indirect action. Then, we tested the effect of HCTZ in WCC currents in HEKcells expressing the BK channel without any β-subunit. In this condition, 100µM HCTZ did not change the BK activity (%current increase (+40 mV): 13.9 ± 15.7,P>0.05; n:4). These experimental results were repeated in HUASMCs.Consistent with the previous results, 10 µM HCTZ caused significant activationof BK current in WCC (528 ± 215 to 1379 ± 132 pA at +40mV, n: 4, p <0.05)while, when it was applied in the IOC, it did not produce any changes in BKchannel open probability (NPo (+40mV): 0.0114 ± 0.0015 (control) vs 0.0135 ±0.0037 (HCTZ), n: 4, P> 0.05). Conclusion:A β1-subunit-dependent mechanism that requires SMC integrityleads to HCTZ-induced BK channel activation.