NOREPINEPHRINE MODULATES DAILY RHYTHMS IN EX VIVO SPLENIC MACROPHAGES

RAMIRES, M; DELGADO, MS; ANZULOVICH AC; CARGNELUTTI E.

Congreso; XXXVII REUNIÓN CIENTÍFICA ANUAL DE LA SOCIEDAD DE BIOLOGÍA DE CUYO; 2019

AbstractThe splenic macrophages (MΦ) phagocytes and eliminates circulatingpathogens, and orchestrate the development of the specific acquired immuneresponse. Even though circadian effects on the immune system have beendocumented, the circadianregulation in the spleen, has not been completely elucidated yet. In mammals, thecentral clock in the suprachiasmatic nucleus (SCN) of the anterior hypothalamussynchronizes cell-autonomous clocks. Communication between SCN and spleenoccurs by the sympathetic nervous system (SNS), through nerves that releasenorepinephrine (NE) in areas of T and MΦ cells. Previously, other authorsreported daily oscillation of NE in spleen. Our focus was to determine whetherrhythmic expression of the molecular clock in the splenic MΦ is regulated bySNS innervation. For this purpose, we disrupted the autonomic innervation tothe spleen of male Holtzman rats, by splenic injections of guanethidine. Animalswere maintained under 12h-light: 12h-dark conditions and ad-libitum food/water intake. First, we evaluated the effects oflocal guanethidine injection on spleen macroscopic appearance, size, weight,and lymphocyte number. Second, to study the NE temporal impact on the molecularclock of splenic MΦ, ten days after guanethidine injection, control andsympathectomized rats were euthanized at different times during a 24 h period(ZT2, ZT6, ZT10, ZT14, ZT18 and ZT22) and spleen was aseptically removed for ex vivo cultures. The clocktranscription factor, BMAL1, was analysed by Western blot, from splenicadherent cells. We found no significant differences in the spleen macroscopicappearance, size, weight, and lymphocyte number, between both treated groups, suggestinglocal guanethidine injections did not have any toxicity on spleen cells. Thesplenic MΦ from control rats showed a circadian oscillation of BMAL1 (% rhythm:71.8), with its acrophase occurring at the middle of the light period. Incontrast, the ex vivo splenic MΦ fromguanethidine-treated animals lost the circadian oscillation of BMAL1. Ourresults would indicate a regulation of the molecular clock in splenic adherentcells by the SCN, through the NE sympathetic pathway.