H+ Inhibits TRIC-B Channels Derived from Mouse TRIC-A Knockout Tissue

Abstract

Trimeric intracellular cation channels (TRIC-A/TRIC-B) are found in the endo/sarcoplasmic reticulum (ER/SR) and nuclear membranes. TRIC-B knockout mice die in respiratory failure following birth but there are additional diverse, severe pathological symptoms (Yamazaki D. et al. (2009)). The physiological roles of TRIC-B are not fully understood and few regulators of channel activity are known. We here investigate if TRIC-B function is sensitive to pH by incorporating light SR membrane vesicles from TRIC-A knockout skeletal muscle into artificial membranes under voltage-clamp conditions in symmetrical 210 mM K-PIPES, pH 7.2. Open probability was determined at ±30 mV when ≤2 channels were gating in the bilayer. With ≥3 channels present, noise analysis was performed because the complex sub-conductance state gating prevented accurate measurements. As previously observed (Venturi E. et al. (2009)), TRIC-B was more open at positive potentials but exhibited variable gating behaviour. For example, at pH 7.2, at +30 mV, noise analysis yielded a mean current of 1.25±0.25 pA (SEM; n=23) but ranged from 0.025 to 3.36 pA. We observed that TRIC-B became more open as cytosolic pH was increased. At +30 mV, as cytosolic pH was raised from 7.2 to 9.2, we observed a 3.34 fold increase in mean current (SEM; n=7; p<0.05) whereas mean current decreased 3.33 fold when cytosolic pH was lowered from pH 7.2 to 6.2, A similar trend was observed when luminal pH was altered. Under acidic conditions where both cytosolic and luminal pH was lowered, TRIC-B channel opening was markedly inhibited and the effects of cytosolic and luminal [H+] appeared to be additive.Further experimentation is required to understand the mechanisms underlying pH regulation of TRIC-B function and the physiological/pathophysiological significance of TRIC-B sensitivity to pH.Supported by the BHF, EPSRC and JSPS.