Excitatory signal transduction mediated by inositol phospholipid metabolism in lobster (Panulirus argus) olfactory receptor neurons

Appropriate primary sustained culture conditions were developed to study signal transduction in Panulirus argus olfactory receptor neurons (ORNs). Neurons were cultured in a modified Liebowitz media supplemented with salts, vitamins, L-glutamine, low dextrose, and either fetal calf serum or lobster haemolymph. The nature of the adequate stimuli, the degree of tuning of the cells, the threshold of sensitivity, and the dual polarity of the odor-evoked currents were consistent with chemosensitivity in the cultured ORNs being olfactory.; The magnitude of the odor-evoked currents was significantly increased or decreased by nonhydrolyzable analogs of GTP and GDP, respectively, and not perturbed by pertussis and cholera toxins, implying that a class of bacterial toxin-insensitive G-proteins was mediating signal transduction. An antibody directed against {dollar}rm Gsb{lcub}oalpha{rcub}{dollar} immunolabelled a 40.5 kDa band in an enriched membrane preparation of ORN outer dendrites and, along with an antibody directed against {dollar}rm Gsb{lcub}qalpha{rcub},{dollar} selectively decreased the odor-evoked inward current within 10 min of initial perfusion.; Inositol 1,4,5-trisphosphate {dollar}rm (IPsb3){dollar} selectively evoked an inward current in the ORNs. Application of {dollar}rm IPsb3{dollar} to the inside face of cell-free patches of ORN plasma membrane directly gated two ion channels that differed in conductance, voltage dependence, and dwell-time kinetics. An antibody directed against an intracellular, cerebellar {dollar}rm IPsb3{dollar} receptor recognized a protein of similar molecular weight to the mammalian receptor in the ORNs and was found to increase selectively the odor-evoked inward currents and {dollar}rm IPsb3{dollar}-activated unitary currents in the lobster ORNs.; Modulation of channel gating or ion permeation was observed in both {dollar}rm IPsb3{dollar}-gated channels in response to elevated pH or {dollar}rmlbrack Casp{lcub}2+{rcub}rbracksb{lcub}i{rcub}.{dollar} Both channels mimicked the pharmacology of the macroscopic odor-evoked inward currents. Ion substitution suggested that the small-conductance channels were nonselective for cations and that the large-conductance channels were either nonselective between {dollar}rm Nasp+{dollar} and {dollar}rm Casp{lcub}2+{rcub}{dollar} or were selective for {dollar}rm Casp{lcub}2+{rcub}.{dollar}; The direct metabolite of {dollar}rm IPsb3,{dollar} inositol 1,3,4,5-tetrakisphosphate {dollar}rm (IPsb4),{dollar} gated an ion channel that differed in conductance, density, kinetics, and voltage sensitivity from those activated by {dollar}rm IPsb3.{dollar} The {dollar}rm IPsb4{dollar}-gated channel mutually interacted with {dollar}rm IPsb3{dollar}-gated channels to alter the open probability of the channels.