TRPM5 is activated by micromolar concentrations of Ca²⁺.(a) Activation by 40 μM Ca²⁺ of an inward current in a patch excised from a TRPM5-transfected CHO-K1 cell (V_m =-80 mV). Neither 10 μMIP₃ nor 100 μM OAG elicited a current in the same patch. (b) Single-channel recording at different voltages in response to 12 μM Ca²⁺ (Left). The I–V relation of the same patch (Right) yields a slope conductance of 16 pS. No channel activity was seen in this patch in the absence of Ca²⁺.(c) Responses of TRPM5 channels to increasing concentrations of Ca²⁺. Note the desensitization of the channels in response to repeated application of the stimulus protocol. (d) Dose–response relations from patches (V_m = -80 mV) before (filled circles) or after (open circles) desensitization (mean ± SEM, n = 3 patches; data were normalized to the peak current in each patch). Fits are to the Hill equation with K_1/2 = 21 μM, n = 2.4, and V_max = 1.05 before desensitization (solid line) and K_1/2 = 77 μM, n = 2.4, and V_max = 0.23 after desensitization (dashed line; note this fit was obtained by holding n constant, but allowing V_max and K_1/2 to vary). The dotted line shows the predicted relationship if only the maximum current were to decline (simulation with K_1/2 = 21 μM, n = 2.4, and V_max = 0.23), illustrating that with desensitization the dose–response relationship shows a shift in sensitivity as well as a decrease in the maximal current activated.

Desensitization of TPRM5 is Ca²⁺-dependent. (a) Responses in an excised patch (-80 mV) to repeated applications of Ca²⁺. Note that the current declined gradually in response to repeated exposures to 40 μM Ca²⁺ but declined dramatically in response to 500 μM Ca²⁺. (b) Response in patches to prolonged exposure to Ca²⁺ (-80 mV). Single exponential fits are shown in red. (c) The rate of decay of the current (1/τ) is linearly related to Ca²⁺ concentration, over the range measured, indicating that desensitization is Ca²⁺-dependent (mean ± SEM, n = 5 patches for each data point).

PIP₂ partially restores TRPM5 channel activity after desensitization. (a) Responses to 12 and 40 μM Ca²⁺ in the presence and absence of 20 μM PIP₂ before and after desensitization. Desensitization was induced by a 30-s exposure to 40 μM Ca²⁺ (V_m = -80 mV). PIP₂ enhances the current in response to Ca²⁺ after, but not before, desensitization. (b) Enhancement of the current in response to 40 μM Ca²⁺ by PIP₂ before (n = 6) and after desensitization (n = 8). Current amplitudes are the averaged peak magnitude recorded within 2 s from the start of Ca²⁺ exposure. The asterisk indicates that the difference between the enhancement of control and desensitized currents was significant at P < 0.05. (c) Dose–response relations before desensitization (open triangles), after desensitization (filled circles), and after desensitization in the presence of 10 μM PIP₂ (open circles) (mean ± SEM, n = 3 patches). In these experiments, PIP₂ was present before and during a 6-s application of Ca²⁺. Currents are normalized to the maximum current obtained in each patch. Solid lines are the same fits as in Fig. 1d. The dashed line shows the expected relationship if 25% of the current recovered full sensitivity. The data could not be well fitted by assuming that either only the maximum current or the sensitivity was restored by PIP₂.

Electrophysiological properties of TRPM5 expressed in HEK-293 M1 cells. (a) Application of the Ca²⁺ ionophore A23187 (20 μM) in the presence of 2 mM extracellular Ca²⁺ in perforated-patch recording mode elicits a large outwardly rectifying current. (Inset) The current in response to a ramp depolarization (1 V/s) at the times indicated. (b) Application of ACh (100 μM) induced a transient current in HEK-293 M1 cells cotransfected with TRPM5 and G16z44. (Inset) The current in response to a ramp depolarization (0.44 V/s) at the times indicated. The pipette solution contained no Ca²⁺ buffer. (c) In whole-cell recording mode, 40 μM Ca²⁺ in the pipette elicited a large rectifying current. Recording began shortly after break into the whole-cell mode. (Inset) The current in response to a ramp depolarization (1 V/s). (d) Currents in response to a family of step depolarizations and the resulting I–V relationship for the peak current at each voltage. Steps are to 0–100 mV from a holding potential of -80 mV with repolarization to -50 mV. Note the prominent relaxation of the current upon depolarization, consistent with voltage-dependent gating of the channels. A small fraction of the current showed a fast increase, indicating that there is low, but nonzero, probability of opening at the resting potential (-80 mV).

A model for taste transduction. Binding of taste stimuli to G protein-coupled taste receptors (R) leads to dissociation of the heterotrimeric G protein. βγ subunits of the G protein activate PLCβ2, which in turn hydrolyzes PIP₂ into DAG and IP₃. IP₃ activates IP₃ receptors, which release Ca²⁺ from intracellular stores. Intracellular Ca²⁺ opens TRPM5 channels, leading to an influx of Na⁺ and depolarization of the cell. Note that TRPM5 is not permeable to Ca²⁺ and, therefore, there is no positive feedback loop.