Imidazoline receptor proteins are regulated in platelet-precursor MEG-01 cells by agonists and antagonists.
The I1-imidazoline receptor is a novel brainstem modulator of sympathetic outflow that is elevated on platelets and in brains of depressed patients. A positive correlation has been reported (accompanying manuscript) between plasma norepinephrine (NE) concentrations and the densities (Bmax) of platelet I1 binding sites (I1 sites). I1-candidate proteins of 33 kDa and 85 kDa are now identified on Western blots probed with anti-imidazoline receptor antiserum (IRBP antiserum), that correlate with Bmax values for I1 sites. Furthermore, a human megakaryoblastoma cell line (MEG-01) has been used to study the regulation of these proteins on megakaryocytic cells, while bovine adrenal chromaffin cells provide a standard I1 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 I2 imidazoline binding sites located on platelet mitochondrial membranes. The 85 kDa band 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-01 cells grown in the presence of 10% FBS; suggesting that a precursor (85 kDa) and product (33 kDa) relationship might be induced by serum. The 85 kDa band was robustly up-regulated in response to imidazoline receptor-sensitive ligands; moxonidine, idazoxan and agmatine (10 microM each for 6 h). NE also up-regulated the 85 kDa IRBP-immunoreactive protein on MEG-01 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-imidazoline alpha 2-antagonist, was ineffective alone, or in combination with moxonidine (up to 40 microM), but yohimbine blocked NE's induction of the 85 kDa band. Therefore, a rise in either plasma NE and/or endogenous I-site ligands (i.e. agmatine) could explain an elevation of imidazoline receptors observed in depression.
Department of Psychiatry and Human Behavior, University of Mississippi Medical Center, Jackson 39216-4505, USA.
T R
TR
Ivanov
Y
Y
Feng
H
H
Wang
S
S
Regunathan
D J
DJ
Reis
D N
DN
Chikkala
P
P
Gupta
J C
JC
Jones
J E
JE
Piletz
D000376:Agmatine / Q000494:pharmacology; D000818:Animals; D001792:Blood Platelets / Q000187:drug effects*; D002417:Cattle; D006801:Humans; D019329:Idazoxan / Q000494:pharmacology; D007093:Imidazoles / Q000494:pharmacology; D054755:Imidazoline Receptors; D009638:Norepinephrine / Q000097:blood; D011955:Receptors, Drug / Q000187:drug effects*; D014407:Tumor Cells, Cultured / Q000187:drug effects*; D015854:Up-Regulation / Q000187:drug effects; D015016:Yohimbine / Q000494:pharmacology
D007093:Imidazoles; D054755:Imidazoline Receptors; D011955:Receptors, Drug; D015016:Yohimbine; D000376:Agmatine; C043482:moxonidine; D009638:Norepinephrine; D019329:Idazoxan