160 Brai~t Regearch, 490 (198~t) 166-1~9 Elscvicl BRE 23560 Twenty-four hour variation of l-adrenergic receptors in the pineal gland of the male Syrian hamster Andreas Pangerl l, Brigitte Pangerl 1, Russel J. Reiter 1, George M. Vaughan 3 and David J. Jones 2 1Departments of Cellular and Structural Biology and 2Anesthesiology, The University of Texas Health Science Center at San Antonio, San Antonio, TX 78284 (U.S.A.) and ~ US Army Institute of Surgical Research, Fort Sam, Houston. TX 78234-5012 (U.S.A.) (Accepted 28 February 1989) Key words: Pineal gland; a~-Adrenoceptor: Circadian variation Specific binding of [~25IJiodo-[fl-(4-hydroxyphenyl)-ethylaminomethyl]tetralone ([125I]HEAT) was used to assess a~-adrenergic receptors on pineal gland membranes of male Syrian hamsters (Mesocricetus auratus) housed under a 14:10 h light-dark cycle (lights on at 06.00 h). Saturation experiments with pooled pineal membrane preparations showed the presence of al-adrenergic receptor sites (dissociation constant K d approx. 0.1 nM). Analysis of 4 time points indicated no significant change in Kd, but significant (P < 0.01) changes of receptor density (Bmax) with a minimum recorded at night. Binding of a constant amount of [125I]HEAT (200 pM) to pineal membranes at 8 time points exhibited a circadian variation (P < 0.001) of receptor density with lowest values around midnight and highest levels during daytime. Melatonin synthesis in the pineal gland of most species is regulated primarily via a fl-adrenergic receptor mechanism. Stimulation of fl-adrenoceptors by norepinephrine or other fl-adrenergic agonists induces an increase of pineal cAMP content, N- acetyltransferase activity (NAT) and consequently of melatonin content 2'13. Both in rat 19 and Syrian hamster 17 the response of the pineal gland to fl-adrenergic stimulation is potentiated by a-adren- ergic agonists. The presence of a-adrenergic receptors in the pineal gland of the rat is well established with the receptor being of the al-type 19. However, a 24 h variation in a-receptor density is found only in the pineal gland of ovariectomized females2°, but not in male rats; conversely, in the rat pineal a 24 h rhythm of pineal fl-adrenergic receptors is well documen- ted7J 4,16. The present study is the first demonstration of al-adrenergic receptors in the pineal gland of the Syrian hamster. Having established the presence of these receptors in the pineal gland of the Syrian hamster, we then explored whether a-receptor num- ber varied over the 24 h period. [125I]HEAT ([12sI]iodo-[fl-(4-hydroxyphenyl)-eth- ylaminomethyl]tetralone, spec. act. 2200 Ci/mmol) was purchased from New England Nuclear (Boston, MA) and stored at -20 °C. Polyethylene glycol (approximate mol. wt. 8000) and Tris were pur- chased from Sigma, St. Louis, MO. Phentolamine was a gift from Ciba Geigy, Summit, NY. Bio Rad Protein Assay Kit 2 was purchased from Bio Rad, Richmond, CA. All other chemicals were analytical grade or better from standard commercial sources. Adult male Syrian hamsters (Mesocricetus au- ratus, 80-90 g) were obtained from Sasco (Omaha, NE) and housed under a daily 14:10 h light-dark regime with lights off at 20.00 h. Animals were decapitated (during the dark period under dim red light), the pineal gland rapidly removed, immedi- Correspondence: R.J. Reiter, Department of Cellular and Structural Biology, The University of Texas Health Science Center at San Antonio, 7703 Floyd Curl Drive, San Antonio, TX 78284, U.S.A.