Activation by bitter substances of a cationic channel in membrane patches excised from the bullfrog taste receptor cell

Abstract

The response to bitter-tasting substances was recorded in outside-out membrane patches excised from the taste receptor cell of the bullfrog fungiform papilla. Application of a bitter-tasting substance, quinine or denatonium, induced channel openings under conditions in which none of the second messenger candidates or their precursors (e.g. cyclic nucleotide, inositol 1,4,5-trisphosphate, Ca2+, ATP and GTP) were present on either side of the membrane. The response could be recorded > 10 min after excision of the patch membrane. These data suggest that the channel was directly gated by the bitter-tasting substances. No change in response was detected upon addition to the cytoplasmic side of either GDPβS (1 mM) or GTPγS (1 mM), suggesting that the G protein cascade has no direct relation to response generation. The quinine-induced current was dose dependent. The lowest effective concentration was approximately 0.1 mM, and the saturating concentration was near 1 mM. The dose-response curve was fitted by the Hill equation with a K½ of 0.52 mM and a Hill coefficient of 3.8. The single channel conductance measured in 120 mM NaCl solution was 10 pS. The channel was cation selective, and the ratio of the permeabilities for Na+, K+ and Cs+ (PNa:PK:PCs) was 1:0.48:0.39. The unitary conductance was dependent on the extracellular Ca2+ concentration ([Ca2+]o); 9.2 pS in a nominally Ca2+-free solution, and 4.5 pS in 1.8 mM [Ca2+]o. The dose dependence, the ion selectivity and the dependence of the unitary conductance on [Ca2+]o were almost identical to those of the quinine-induced whole-cell current reported previously, indicating that the channel activity observed in the excised membrane is the basis of the whole-cell current. The present observations suggest the new possibility that the cationic channel directly gated by bitter substances is involved in the bitter taste transduction mechanism.