Effect Of Estradiol On The Expression Of Serotonin And Serotonin 1A And 2A Receptor Subtypes In Ovariectomized Rats

OBJECTIVE: Hot flushes (flashes in USA) are the most common complaint of perimenopausal and postmenopausal women. The pathophysiology of hot flushes remains unknown, which limits valid control of hot flushes. The previous viewpoint thought that a decline in hormone concentrations might lead to hot flushes. Estrogen Replacement Therapy (ERT) has been used for a long time to treat menopausal symptom, including hot flushes. It also be seemed to be the most effective treatment for hot flushes. However, many women are reluctant to accept ERT for fear of cancer and the existence of ERT contraindication. Recently, the benefits and the risk of long-term ERT are still to be questioned. Therefore, the search for other non-hormonal treatments to alleviate hot flushes symptom and improve life quality in women has generated an enormous amount of interest. Unfortunately, such treatments are not very effective and often associated with adverse effects. Results from recent studies showed that selective serotonin(5-HT)reuptake inhibitors (SSRIs) and other similar compounds can safely and effectively reduce hot flushes. These results suggest that serotonin may be closely related to menopausal hot flushes.But which serotonin receptor subtypes may play key roles in the occurrence of hot flushes are still not known. Studies from pharmacology indicated that serotonin 1A and 2A receptor subtypes have taken part in thermoregulation, but lack of morphological evidence of these two receptors subtypes. It has been repeatedly demonstrated that the hypothalamus is intensely innervated by 5-hydroxytriptamine(5-HT) fibers arising from the raphe nucleus. 5-HT afferent fibers innervate synaptically the hypothalamic neurosecretory neurons controlling their functional activity, such as thermoregulation. Therefore, in present study, we examined the effect of estradiol on the expression of serotonin and serotonin 1A and 2A receptor subtypes in ovariectomized rat.This may facilitate the understanding the mechanism of hot flushes and the search for strategy of coping with hot flushes in non-hormonal drugs.METHODS: Constrution of ovariectomized rats model,immunohistoch-emistry and image analysis technique were used to investigate the effect of estradiol on the change of expression of 5-HT immunoreactivities (5-HT IR) in rapheal nuclei complex of ovariectomized rat. And the same technique was used to observe the distribution pattern of serotonin 1A receptor subtype (5-HTiaR) and 5-HT2aR immunoreactivity in the rat hypothalamus . The specific oligonucleotide primers were synthesized based on the complementary DNA sequencs for the rat 5-HTia and 5-HT2A receptor subtypes. The method of reverse transcription polymerase chain reaction (RT-PCR) was used to detect the presence of 5-HTia and 5-HT2A receptor subtypes in ovariectomy rat hypothalamus after accepted ERT for different periods of time(3 d, 6 d, 9 d, 12 d),which may be helpful to elucidate the pathophysiology of hot flushes. RESULTS: The immunohistochemical results showed that: (1) 5-HT IR was expressed significantly in the nucleus raphe dorsalis and median nucleus of rats. Ovariectomized group caused a decline in cell numbers and optical denisity(OD) of 5-HT IR in both sites, compared with Sham ovariectomized control group(p<0.05). Ovariectomized supplying E2 group is similar to the sham rats (P<0.05). (2) Intensely and densely stained 5-HT_1aR immunoreactive neurons were observed in magnocellular nucleus of preoptic area, preoptic periventricular nucleus, supraoptic nucleus and lateroanterior hypothalamic nucleus. Considerably weaker immunoreactivity was detected in medial preoptic nucleus, lateral preoptic area, periventricular hypothalamic nucleus, lateral hypothalamic area, dorsomedial hypothalamic nucleus, ventromedial hypothalamic nucleus, tuber cinereum, tuberomammillary nucleus, medial mammillary nucleus and lateral mammillary nucleus. Very weak immunoreactivity was found in the median preopitc nucleus, suprachiasmatic nucleus, paraventricular hypothalamic nucleus, dorsal hypothalamic nucleus, arcuate nucleus, posterior hypothalamic area, supramammillary nucleus and premammillary nucleus. (3) 5-HT_2aR immunohistochemically stained products were mainly observed in the fibers and terminals. 5-HT(2a)R immunoreactive cell bodies were mainly present in paraventricular hypothalamic nucleus, supraoptic nucleus, ventromedial hypothalamic nucleus and tuber cinereum, medial preoptic area, lateral hypothalamic area, lateroanterior hypothalamic nucleus and dorsomedial hypothalamic nucleus.The RT-PCR results: (1) The presence of 5-HT1AR and 5-HT2AR mRNA was detected in rat hypothalamus. (2) The expression levels of 5-HTiaR mRNA in 3 d, 6 d OVX supplying E2 (OVX+E2) group and 3 d,...