Modulation of nicotinic acetylcholine receptor channel by pH: a difference in pH sensitivity of Torpedo and mouse receptors expressed in Xenopus oocytes.

1. In this study, effects of pH on the ion channel function of nicotinic acetylcholine receptor (nAChR) from Torpedo californica electroplax and mouse muscle BC3H-1 cells were investigated using Xenopus laevis oocytes injected with in vitro synthesized RNA transcripts. The acetylcholine (ACh)-induced whole-cell peak current responses and slow desensitization rates were measured by voltage-clamp. 2. The ACh-induced peak currents of Torpedo nAChRs were reversibly diminished when extracellular pH was reduced from 7.4 to 5.0 and increased at pH greater than 7.4. This pH dependence had an apparent pKa of 7.0. In contrast, the peak current of mouse muscle nAChRs were reversibly decreased at both acidic and alkaline pH's. This bell-shaped pH profile with a maximum at pH 7.4 had two apparent pKa values of 5.6 and 9.2. 3. The peak current responses of four mouse-Torpedo nAChR hybrids consisting of three Torpedo subunits and one mouse nAChR subunit displayed similar pH profiles at acidic pH, with a pKa of 6.0 to 6.5, which lay between the pKa 5.6 for mouse and the pKa 7.0 for Torpedo nAChRs. Two of these combinations, alpha M beta T gamma T delta T and alpha T beta T gamma M delta T, also had an alkaline pH dependence of the current response similar to that of mouse receptor, with a pKa of 9.3 to 9.5. 4. The slow desensitization rate of Torpedo nAChRs increased at both acidic and alkaline pH's, with two pKa values of 6.5 and 9.5, whereas that of mouse muscle receptors remained unchanged from pH 6.5 to pH 9.0 and increased at pH less than 6.0, with a pKa of 4.7. 5. Substitution of each subunit of Torpedo nAChR with a mouse counterpart resulted in a pH dependence pattern (pKa 6.0 to 6.4 and pKa 9.1 to 9.3) similar to that of Torpedo nAChR except the substitution with mouse beta subunit (pKa 4.8 and pKa 8.9), which appears to carry the characteristic acidic group determining the pH dependence of mouse nAChR desensitization. 6. The apparent pKa values obtained from the pH dependence studies of nAChR channel activation and desensitization suggest the involvement of different amino acid residues in determining channel functions of Torpedo and mouse nAChRs. The groups with pKa 7.0 and 6.5 on Torpedo nAChR can be tentatively identified as His residues, whereas those with pKa 5.6 and 4.7 on mouse receptor as Glu or Asp residues. The alkaline pKa of 8.9 to 9.5 may be Cys, Tyr, or Lys.