Analysis of single cyclic nucleotide-gated channels in olfactory receptor cells

Abstract

In the accompanying article (Firestein et al., 1991b), we have demonstrated that odor- and cyclic nucleotide-sensitive channels exist at a low density in the dendritic membranes of isolated salamander olfactory receptor neurons. Here, we analyze the cyclic nucleotide sensitivity of these channels using the inside-out patch recording technique. Both cAMP and cGMP, at micromolar concentrations, are capable of inducing channel openings. The biophysical parameters of channel activity are nearly the same in response to either ligand. The unitary conductance is about 45 pS, the reversal potential of single-channel currents is +5 mV, and the I/V relation is linear over the range -80 to +80 mV. The channel activity shows no obvious voltage dependence in divalent cation-free symmetrical solutions. The channel shows no desensitization, even to agonist exposures lasting 15 sec. Mean open time is about 1.5 msec; the closed time distribution is best fit by two exponentials with a fast time constant in the submillisecond range (ca. 0.15 msec) and a slower time constant in the millisecond range (ca. 1.5 msec). The only clear difference in the activity of the two ligands is in their affinity constants. The K1/2 for cAMP is 20 microM; that for cGMP is 4 microM. In both cases, the Hill coefficient is greater than 2, suggesting that channel opening requires the cooperative action of three ligand molecules.