IMIDAZOLINE RECEPTOR PROTEINS ARE REGULATED IN PLATELET-PRECURSOR MEG-01 CELLS BY AGONISTS AND ANTAGONISTS

The I-1-imidazoline receptor is a novel brainstem modulator of sympath etic outflow that is elevated on platelets and in brains of depressed patients. A positive correlation has been reported (accompanying manus cript) between plasma norepinephrine (NE) concentrations and the densi ties (B-max) of platelet I-1 binding sites (I-1 sites). I-1-candidate proteins of 33 kDa and 85 kDa are now identified on Western blots prob ed with anti-imidazoline receptor antiserum (IRBP antiserum), that cor relate with B-max values for I-1 sites. Furthermore, a human megakaryo blastoma cell line (MEG-OI) has been used to study the regulation of t hese proteins on megakaryocytic cells, while bovine adrenal chromaffin cells provide a standard I, cell type for comparison. Both the 33 kDa and 85 kDa IRBP-immunoreactive bands were enriched in plasma membrane fractions. IRBP antiserum did not cross-react with I, imidazoline bin ding sites located on platelet mitochondrial membranes. The 85 kDa ban d was enhanced under conditions lacking fetal bovine serum (FBS) from the culture medium 6 h prior to harvesting. Conversely, 33 kDa protein was enhanced on MEG-OI cells grown in the presence of 10%FBS; suggest ing that a precursor (85 kDa) and product (33 kDa) relationship might be induced by serum. The 85 kDa band was robustly up-regulated in resp onse to imidazoline receptor-sensitive ligands; moxonidine, idazoxan a nd agmatine(10 mu M each for 6 h). NE also up-regulated the 85 kDa IRB P-immunoreactive protein on MEG-OI membranes, but to a lesser extent. Idazoxan, an imidazoline alpha(2)-antagonist, off-set its induction of 85 kDa protein by reducing the 33 kDa band. Yohimbine, a non-imidazol ine alpha(2)-antagonist, was ineffective alone, or in combination with moxonidine (up to 40 mu M): but yohimbine blocked NE's induction of t he 85 kDa band. Therefore. a rise in either plasma NE and/or endogenou s I-site ligands (i.e, agmatine) could explain an elevation of imidazo line receptors observed in depression. (C) 1998 Elsevier Science Ltd. All rights reserved.