Ovarian steroids decrease DNA fragmentation in the serotonin neurons of non-injured rhesus macaques.

Abstract

We previously found that ovarian steroids promote neuroprotection in serotonin neurons by decreasing the expression of pro-apoptotic genes and proteins in the dorsal raphe nucleus of rhesus macaques, even in the absence of overt injury. In this study, we questioned whether these actions would lead to a reduction in DNA fragmentation in serotonin neurons. Ovariectomized (OVX) rhesus monkeys received Silastic implants that were empty (placebo) or containing estradiol (E), progesterone (P) or estradiol plus progesterone (E+P) for one month. Eight levels of the dorsal raphe nucleus in a rostral to caudal direction were immunostained with TUNEL (terminal deoxynucleotidyl transferase nick end labeling). Two staining patterns were observed, which are referred to as type I, with complete dark staining of the nucleus, and type II, with peripheral staining in the perinuclear area. A montage of the dorsal raphe was created at each level with a Marianas Stereology Microscope and Slidebook 4.2 and TUNEL positive cells were counted. In direct comparison with OVX animals, P treatment and E+P treatment significantly reduced the total number of TUNEL positive cells (Mann Whitney test, both treatments p=0.04) and E+P treatment reduced the number of TUNEL positive cells/cubic millimeter (Mann Whitney test, p=0.04). Double immunocytochemistry for TUNEL and TPH indicated that DNA fragmentation was prominent in serotonin neurons. These data suggest that in the absence of ovarian steroids, a cascade of gene and protein expression leads to an increase in DNA fragmentation in serotonin neurons. Conversely, ovarian steroids have a neuroprotective role in the non-injured brain and prevent DNA fragmentation and cell death in serotonin neurons of nonhuman primates.