Abstract

We studied the opening mechanism of Ca²⁺-permeable channels formed with mouse transient receptor potential type 5 (mTRP5) usingXenopus oocytes. After stimulation of coexpressed muscarinic M₁ receptors with acetylcholine (ACh) in a Ca²⁺-free solution, switching to 2 mM Ca²⁺-containing solution evoked a large Cl⁻current, which reflects the opening of endogenous Ca²⁺-dependent Cl⁻ channels following Ca²⁺ entry through the expressed channels. The ACh-evoked response was not affected by a depletion of Ca²⁺ store with thapsigargin but was inhibited by preinjection of antisense oligodeoxynucleotides (ODNs) to G_q, G₁₁, or both. The mTRP5 channel response was also induced by a direct activation of G proteins with injection of guanosine 5′-3-O-(thio)triphosphate (GTPγS). The ACh- and GTPγS-evoked responses were inhibited by either pretreatment with a phospholipase C inhibitor, U73122, or an inositol-1,4,5-trisphosphate (IP₃) receptor inhibitor, xestospongin C (XeC). An activation of IP₃ receptors with injection of adenophostin A (AdA) evoked the mTRP5 channel response in a dose-dependent manner. The AdA-evoked response was not suppressed by preinjection of antisense ODNs to G_q/11 or U73122 but was suppressed by either preinjection of XeC or a peptide mimicking the IP₃ binding domain of Xenopus IP₃ receptor. These findings suggest that the activation of IP₃ receptor is essential for the opening of mTRP5 channels, and that neither G proteins, phosphoinositide metabolism, nor depletion of the Ca²⁺ store directly modifies the IP₃receptor-linked opening of mTRP5 channels.