1. The nature of the binding site mediating the
insulin secretagogue activity of certain
imidazoline compounds remains unclear and the pharmacology of the I1- and I2-imidazoline sites, described in many tissues, does not correlate with the observed responses to
imidazolines in islets. In the present paper, we describe further results which support the concept that the islet
imidazoline site may represent a novel subtype of
imidazoline receptor. 2. Culture of rat isolated islets in the presence of
imidazoline secretagogues (either
efaroxan or
phentolamine) resulted in loss of responsiveness on subsequent re-exposure to these agents. However, culture of islets with either
idazoxan or UK14,304 (
imidazoline ligands that do not stimulate insulin secretion) did not lead to any loss of response when the islets were subsequently exposed to
efaroxan. By contrast, islets cultured with UK14,304 (a potent alpha 2-
adrenoceptor agonist), displayed loss of sensitivity to
noradrenaline, consistent with down-regulation of alpha 2-adrenoceptors. 3. In order to characterize the
imidazoline site further, radioligand binding studies were performed in membranes from RINm5F
insulinoma cells using [3H]-
RX821002, an
imidazoline insulin secretagogue that does not interact significantly with
imidazoline sites in other tissues. [3H]-
RX821002 labelled alpha 2-adrenoceptors with high affinity (2.01 +/- 0.7 nM) but also labelled a second, non-
adrenoceptor site with much lower affinity. 4. Under conditions of alpha 2-adrenoceptor blockade (in the presence of
adrenaline),
efaroxan displaced [3H]-
RX821002 binding to the low affinity site, in a dose-dependent manner. Competition studies employing additional
imidazoline compounds of varying
secretagogue activity revealed that the pharmacological profile of the low affinity site correlates well with that observed in secretion experiments.5. The results obtained from the down-regulation experiments with isolated islets and from the radioligand binding studies suggest that the low affinity [3H]-
RX821002 binding site may represent the functional receptor responsible for the
secretagogue activity of
imidazoline compounds in the endocrine pancreas and that it has a pharmacological profile distinct from those of I,- and 12-sites.