Teramoto Noriyoshi, Tomoda Toshihisa, Yunoki Takakazu, Brading Alison F, Ito Yushi
Department of Pharmacology, Graduate School of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi Ward, 812-8582, Fukuoka, Japan.
Life Sci. 2002 Dec 20;72(4-5):475-85. doi: 10.1016/s0024-3205(02)02284-1.
Patch-clamp experiments have been performed to investigate the effects of endoproteases (such as trypsin, carboxypeptidase B) on both membrane currents and unitary currents in isolated smooth muscle cells from pig proximal urethra (conventional whole-cell configuration, cell-attached configuration, and inside-out patches). Application of either trypsin (1 mg/mL) or carboxypeptidase B (0.1 mg/mL) to the intracellular surface of the excised membrane patches stimulated the activity of a 2.1 pA K+ channel (in symmetrical 140 mM K+ conditions) at a holding potential of -50 mV. The trypsin-induced K+ channels in inside-out configuration exhibited the same amplitude and similar channel opening kinetics to the levcromakalim-induced ATP-sensitive K+ channel (i.e. K ATP channel) in cell-attached patches of the same membrane; however, the sensitivity of the channels to glibenclamide was greatly reduced after the trypsin-treatment. The activity of the trypsin-induced K+ channel was reversibly inhibited by cibenzoline in an inside-out configuration (Ki = 5 microM). It is concluded that trypsin and carboxypeptidase B reactivate the channel with an intact pore activity but the different pharmacological properties of the channels may reflect some change in the conformation in channel proteins after proteolysis.
