The development of physiologic responsiveness to muscarinic agonists in chick embryo heart cell cultures. Role of high affinity receptors and sensitivity to guanine nucleotides.

Prior to ingrowth of the vagus nerve (4-5 days in ovo), embryonic chick hearts are relatively unresponsive to muscarinic stimulation ( Pappano , A. J. (1977) Pharmacol. Rev. 29, 3-33). We studied the correlation between the development of physiologic responsiveness in the embryonic chick heart and changes in the properties of muscarinic receptors. In cultures from hearts 10 days in ovo, muscarinic agonists decreased beating rate by 15% and increased the rate of K+ efflux by 35%. In cultures of embryonic hearts 3 1/2 days in ovo, muscarinic receptors had no effect on beating rate and mediated only an 11% increase in the rate of K+ efflux. We previously demonstrated that in cells cultured from hearts 10 days in ovo, 26% of receptors bound agonist with a high affinity (RH) and that incubation with guanine nucleotides mediated the conversion of RH to a low affinity form (RL ( Galper , J. B., Dziekan , L. C., O' Hara , D. S., and Smith, T. W. (1982) J. Biol. Chem. 237, 10344-10356]. In cultures of hearts 3 1/2 days in ovo, RH constituted 52% of total 3H quinuclidinyl benzilate-binding sites, and guanine nucleotides had no effect on the conversion of RH to RL. Growth of cells cultured from hearts 3 1/2 days in ovo in medium supplemented with specific lots of serum resulted in a 3-fold increase (from 11 to 30%) in the ability of muscarinic agonists to increase K+ permeability. This increased sensitivity to muscarinic stimulation was accompanied by a 20% increase in RH and sensitivity of 75% of RH to guanine nucleotides. Thus, enhanced number of RH and development of guanine nucleotide responsiveness are associated with the development of a physiologic response. The relationship of these developmental changes to the appearance of a guanine nucleotide regulatory protein is discussed.