Imidazoline Receptor Ligands Research Articles

Chapter 19 Metabolism of agmatine (clonidine-displacing substance) by diamine oxidase and the possible implications for studies of imidazoline receptors

Clonidine-displacing substance, thought to be the endogenous ligand for imidazoline receptors, has been identified recently as agmatine (1-amino-4-guanidinobutane). The similarity of this compound's structure to that of the diamine oxidase (DAO) inhibitor, aminoguanidine, led us to investigate the possibility that agmatine might be a substrate for this enzyme. The metabolism of agmatine by purified porcine kidney DAO was measured by a peroxidase-linked colorimetric assay. Agmatine was a substrate for this enzyme and, under the experimental conditions used here, was metabolised at a rate of 0.8 mumol agmatine h-1 (unit DAO activity)-1. In contrast, agmatine was a substrate neither for rat brain monoamine oxidase (MAO) -A or -B, nor for rat brown adipose tissue semicarbazide-sensitive amine oxidase (SSAO). The metabolism of agmatine by DAO was inhibited by aminoguanidine (IC50 14.9 nM) and by the antidepressant, phenelzine (IC50 1.95 microM). These results suggest that administration of DAO inhibitors may increase endogenous agmatine levels and thus alter imidazoline receptor densities. A review of the literature documenting ligand affinities for idazoxan-preferring (I2) imidazoline binding site subtypes and drug affinities for DAO enzymes indicates that some of the I2 sites described elsewhere may correspond to DAO and not to an imidazoline receptor.

Agmatine: an endogenous clonidine-displacing substance in the brain.

Clonidine, an antihypertensive drug, binds to alpha 2-adrenergic and imidazoline receptors. The endogenous ligand for imidazoline receptors may be a clonidine-displacing substance, a small molecule isolated from bovine brain. This clonidine-displacing substance was purified and determined by mass spectroscopy to be agmatine (decarboxylated arginine), heretofore not detected in brain. Agmatine binds to alpha 2-adrenergic and imidazoline receptors and stimulates release of catecholamines from adrenal chromaffin cells. Its biosynthetic enzyme, arginine decarboxylase, is present in brain. Agmatine, locally synthesized, is an endogenous agonist at imidazoline receptors, a noncatecholamine ligand at alpha 2-adrenergic receptors and may act as a neurotransmitter.

Imidazoline receptors in rat liver cells: A novel receptor or a subtype of α2-adrenoceptors?

An imidazoline/guanidine receptor has been characterized in rat liver cells. Binding of [ 3H]idazoxan, a selective benzodioxan antagonist, to imidazoline receptor on intact fresh hepatocytes (B max = 801 ± 23 fmol/mg protein,K d = 11 ± 0.8 nM) and to liver membranes (B max = 400 ± 38 fmol/mg protein, K d = 10 ± 2 nM) was saturable at 4°C within 3.5 h and at 30°C within 30 min, respectively. Rat lung membranes had more imidazoline sites (B max = 578 ± 30 fmol/mg protein, K d = 14 ± 1.4 nM) than α 2-adrenoceptors (B max = 175.0 ± 20.0 fmol/mg protein,K d = 4.8 ± 2.0 nM). We also screened other tissues for imidazoline sites; labeled with [ 3H]idazoxan displaced by cirazoline was lower in rat lung compared to rat brain and human platelets. The imidazoline receptor has common pharmacological properties with α 2-adrenoceptors, although it is not a subtype of the adrenoceptor, since it bound neither the endogenous agonists norepinephrine and epinephrine, nor the selective α 2-antagonists yohimbine and phentolamine. All guanidine type α 2-adrenoceptor drugs (e.g. guanabenz, guanoxan) and imidazolines (e.g., UK-14,304, naphazoline) competed with high affinity for the liver imidazoline receptor. The lack of effect by Gpp(NH)p, a non-hydrolysable GTP analogue, on the affinity of guanidine- and imidazoline-type ligands for liver imidazoline receptors suggests that the mode of action of these drugs at imidazoline receptors is different than at conventional α 2-adrenoceptors. Ionic changes were considered as a possible mechanism underlying the α 2-adrenoceptor effects in various cells. Opening of K + channels by α 2-adrenoceptors agonists is a pathway which might be shared by imidazoline-type agonists at imidazoline sites. Indeed, 4-aminopyridine, a K + channel blocker, inhibited the specific binding of [ 3H]idazoxan to liver cells with an IC 50 of 0.34 ± 0.07 mM, a concentration which is effective in blocking K + channels in neuronal cells. Similarly, Cs + and NH 4 + effectively interfered with [ 3H]idazoxan binding, suggesting a possible coupling of imidazoline sites to K + gating. The endogenous ligand clonidine-displacing substance (CDS), which was isolated from bovine brain and which binds to α 2-adrenoceptors in brain membranes and human platelets competed with idazoxan at rat liver imidazoline receptors. Although the need of an endogenous ligand for imidazoline sites is obvious, it has yet to be determined whether CDS is the endogenous ligand for imidazoline receptors in general, and for liver cells in particular, and whether K + channels are involved in the mechanism of action of these sites.