Neuronal and inducible nitric oxide synthase in the HPA axis activation by interleukin -1β

J TADEUSZ, A GADEK-MICHALSKA, P RACHWALSKA, J SPYRKA, J BUGAJSKI.

Institute of Pharmacology Polish Academy of Science, Departmentof Physiology 31-343, Poland

In the present study the involvement of neuronal (nNOS) and inducible (iNOS) nitric oxide synthase in the interleukin 1β (IL-1β)-induced activation of hypothalamic-pituitary-adrenal (HPA) axis was determined. Experiments were performed on male Wistar rats (6 weeks old, 190-220 g). Rats were housed in groups of 5 per cage, under controlled conditions (12 h light/dark cycle; a constant temperature 22±2°C) for one week. Groups of rats (n=6-8) were injected i.p.IL -1β (1μg/rat) or IL -1β receptor antagonist (IL-1 ra) (50 μg/kg) 15 min before IL -1β. After decapitation 1, 2 or 3 h after IL -1β injection trunk blood for plasma determinations was collected, the brain was removed from the skull and prefrontal cortex, hippocampus and hypothalamus were excised on a cold plate and snap frozen. Total ACTH, corticosterone and IL -1β levels were determined in plasma. Western blot analyses were performed and IL -1β, nNOS and iNOS were determined in prefrontal cortex, hippocampus and hypothalamus samples.

IL -1β given i.p. induced considerable increase in IL -1β content in prefrontal cortex, hippocampus and hypothalamus 1 h after administration, a strong reduction of this increase appeared after 2 h and a sharp return to high levels 3 h after its administration.

Exogenous IL -1β induced the most pronounced increase of nNOS content in hypothalamus and prefrontal cortex 1 h after administration which gradually decreased 2 h and 3 h later. In the hippocampus nNOS level gradually increased 2 and 3 h after IL -1β administration.

Exogenous IL -1β induced the strongest increase of iNOS in prefrontal cortex and hypothalamus and a weak increase in hippocampus 1 h after administration.

IL -1β i.p. induced a regular, continuous increase of iNOS in hypothalamus to a highest level 3 h after administration. No marked alterations of iNOS content appeared in hippocampus.

Both nNOS and iNOS significantly and to a similar extent increased 1 h after IL -1β administration and gradually declined in a parallel manner 2 and 3 h later.

Much stronger increase of IL -1β content than nNOS and iNOS levels appeared in prefrontal cortex 1 h following IL -1β i.p.

Moderate alterations in nNOS and iNOS levels in hippocampus did not parallel very strong increase of IL -1β content in that brain structure. The highest level of nNOS in hypothalamus 1 h after i.p. IL -1β which regularly declined 2 and 3 h later, contrasted the opposite regular linear increase of iNOS in that structure. The decrease of nNOS was parallel to a similar decline of IL -1β content in the hypothalamus.

IL -1β i.p. increased considerably its plasma level parallel to ACTH level, the highest level 1 h after IL -1β injection. Plasma corticosterone reached the highest level 2 h after IL -1β injection due to both immune and neurohormonal activation of its stimulatory mechanisms in the adrenal cortex.

Pretreatment with IL -1β receptor antagonist reduced the i.p. IL -1β-induced nNOS and iNOS levels in prefrontal cortex, hypothalamus and hippocampus. This antagonist also decreased plasma IL -1β as well as ACTH and corticosterone levels.

These results suggest the involvement of IL -1β, nNOS and iNOS in brain structures, particularly prefrontal cortex and hypothalamus, in regulation of plasma IL -1β, ACTH and, to the same extent, corticosterone secretion under basal conditions.

This research was supported by a grant POIG 01.01.02-12-004/09-00 ”Depression-Mechanisms-Therapy” financed by European Regional Development Fund.