Selective masking of M1-receptors in calf retina membranes by the venom of the marine snail Conus tessulatus

The non discriminatory antagonist [ 3H]QNB labels M1- and M2-muscarinic receptors in calf retina membranes. Crude venom from the marine gastropod Conus tessulatus produces a partial decrease in [ 3H]QNB binding. The total number of sites (560 ± 13 fmol/mg protein in control experiments) decreases to 370 ± 10 fmol/mg protein whereas the affinity of the radioligand is unaffected ( K D = 0.42 ± 0.01 nM and 0.46 ± 0.02 nM, respectively). This process is venom concentration-dependent, quasi-irreversible, and calcium-dependent. Proteolytic activity can not be detected. The partial effect of the venom is related to preferential masking of the M1-receptors. Competition curves of the M1-selective antagonist pirenzepine are shallow in control experiments: 45% of the receptors are of the M1-type ( K i = 45 ± 6 nM) while the remaining are of the M2-type ( K i = 1.0 ± 0.2 μM). In venom-treated membranes, only a low affinity site (M2-receptors, K i = 1.5 ± 0.4 μM) is detected by pirenzepine competition binding. Saturation binding experiments reveal that the venom causes a substantial decrease in the number of high affinity sites for [ 3H]pirenzepine without affecting its K D (23 ± 4 nM and 20 ± 6 nM in control- and venom-treated membranes respectively). The venom produces a leftward shift of the carbachol/[ 3H]QNB competition binding curve, but the ability of 0.1 mM GTP to confer a rightward shift of the competition curve is not affected. These data suggest that the venom from Conus tessulatus contains one or more components which specifically shield part of the M1-receptors (75% in calf retina membranes), leaving the M2-receptors and their functional coupling to G proteins unaffected.