Novel clonidine binding sites were characterized in the intestinal membrane isolated from seawater eels. The specific clonidine binding sites consisted of at least two classes, high affinity (Kd = 1.4 ± 0.3 nM, n = 5) and low affinity (Kd = 175 ± 34 nM, n = 5) sites. The specific binding of 2 nM [3H]clonidine was most enhanced at 20°C and at pH 7.5, and reversed by unlabelled clonidine. Such binding was hardly inhibited by adrenaline, yohimbine or rauwolscine, indicating that most binding sites are distinct from α2-adrenoceptor. The specific clonidine binding was inhibited by various imidazoline/guanidinium drugs, indicating existence of imidazoline/guanidinium receptive sites (IGRS) or imidazoline receptors in the eel intestine. Competition experiments revealed that rank order to displace 2 nM [3H]clonidine from their binding sites was as follows: guanabenz > cirazoline = naphazoline = UK14304 = ST587 ≥ clonidine ≥ idazoxan = RX821002 = tolazoline > ST93 = oxymetazoline = amiloride = ST91 > yohimbine = efaroxan = rauwolscine ≥ adrenaline = ST567 = histamine = agmatine. The rank order was different from those in I1 or I2 sites of IGRS reported in various mammalian tissues, suggesting existence of new IGRS, non I1 and non I2 sites, in the eel intestine. In addition, structure-affinity relationships are discussed from the results of competition experiments. Although physiological role of IGRS is not clear yet even in mammalian cells/tissues, eel intestine may be a good model to elucidate how the IGRS act in the cell and to decide what is the endogenous ligand for the IGRS, since eel intestine contains great amount of IGRS and it responds to guanabenz, an exogenous clonidine derivative.
ASJC Scopus subject areas
- Animal Science and Zoology