(A) Calcium flux elicited by CFA (2.3 mg/ml per 180
μg/cm2), icilin (50
µM), and menthol (500 µM)
on GCaMP6-overexpressing cells transiently transfected with either human
TRPM8 wild type (WT; n = 47, 16, and 47 for icilin,
menthol, and CFA, respectively) or the TRPM8 icilin and menthol binding site
mutant Y745H (n = 3, 3, and 3 for icilin, menthol, and CFA,
respectively). (B) Calcium flux elicited by CFA (2.3 mg/ml per 180
μg/cm2), icilin (50
µM), and menthol (500 µM)
on GCaMP6-overexpressing cells transiently transfected with human TRPM8-WT
(n = 47, 16, and 48 for icilin, menthol, and CFA,
respectively), the PIP2 binding site mutant R1008Q
(n = 3, 3, and 3 for icilin, menthol, and CFA,
respectively), or adjacent residue L1009R (n = 3, 3, and 3
for icilin, menthol, and CFA, respectively). Calcium flux responses were
first subtracted for control vector (CV) responses and then normalized to
responses of the TRPM8-WT cells. Values represent the mean ± S.E.M.
Statistically significant responses using two-way analysis of variance and
post-testing using the Bonferroni multiple comparisons test relative to
TRPM8-WT are represented as *P < 0.05 and
***P < 0.001.