Study Of The Distribution Of Serotonin 5-HT_2A, 5-HT₇ Receptors And Galanin In Spinal Onuf's Nucleus In Rats

Objective: To investigate the distribution of serotonin 5-HT_2A, 5-HT₇ receptors and galanin (GAL) in different motoneuron populations in spinal Onuf's nucleus of rats, and their relationship with the central modulation of the urogenital reflexes.Methods : Twenty-four adult SD rats (twenty-two male rats and two female rats) were randomly divided into gender control group (n=6, include four male and two female rats) , HRP group (n=6) and PRV group (n=12). In gender control group, after the spinal L5-6 segments were removed and cut, sections were treated with 5-HT_2A, 5-HT₇ receptors and GAL immunohistochemical stain, respectively. In HRP group, HRP was injected into right ischiocavernosus muscles (HRP-IC group; n=3) or right external urethral sphincter (HRP-EUS group; n=3), respectively. Twenty-four hours later, the spinal L5-6 segments were cut off and sections were prepared with vibratome. After being treated with TMB method,the sections were treated with 5-HT_2A or 5-HT₇ receptors immuohistochemical detection. Sections were observed by light microscopy and electron microscopy to detect the distribution of 5-HT_2A,5-HT₇ receptors in Onuf's nucleus. In PRV group, recombinant pseudorabies virus (PRV-152 ) were injected into right ischiocavernosus muscles (PRV-IC group; n=6) or right external urethral sphincter (PRV-EUS group; n=6),respectively. Four days later, the spinal L5-6 segments were cut off and sections were prepared with vibratom. Sections were treated with 5-HT_2A or 5-HT₇ receptors or GAL double-labeling immunofluorescence stain and observed by fluorescence microscopy or the laser confocal microscopy to detect the distribution of 5-HT_2A,5-HT₇ receptors and GAL in Onuf's nucleus .Results: In gender control group, 5-HT_2A and 5-HT₇ receptors and GAL expression of male rats in Onuf's nucleus were more dense than female rats; 5-HT_2A receptors immunoreactivity mainly located in the medial part of the dorsolateral nucleus (DLN) of Onuf's nucleus, and 5-HT₇ receptors mainly located in the lateral part of the DLN, however, GAL immuoreactivity located in the medial and lateral part of the DLN. In HRP-IC group, peroxidase labeled motoneurons were located in the medial part of the DLN, in the same area where 5-HT_2A receptor immunoreactivity was found; In HRP-EUS group, HRP labeled motoneurons located in the lateral part of the DLN, 5-HT₇ receptor immunoreactivity overlapped with HRP labeled EUS motoneurons. At the ultrastrutural level, the enzymatic product appeared as needle crystals, immunoreactivity against 5-HT_2A receptors was mainly located in plasma membrane of motoneurons innervating the IC muscle and dendrites surrounding these motoneurons in males rat. However, the 5-HT₇ receptor was mainly localized in subcellular organelles such as mitochondria,the endoplasmic reticulum and Golgi apparatus innervating the EUS in males. In PRV group, after PRV-152 injection in the IC, the PRV retrogradely labeled motoneurons were mainly found in the dorsomedial part of the ispilateral DLN. After PRV-152 injection in the EUS, PRV labeled motoneurons were found ispilaterally in the ventrolateral part of the ispilateral DLN. Immunofluorescence double-labeling detection exhibited that 5-HT_2A receptors mainly distributed in surrounding motoneurons and processes innervating the IC muscles , 5-HT₇ receptors mainly expressed in neurons innervating the EUS,and GAL immunolabeling located in the motoneurons of the EUS and the IC.Conclusion: 5-HT_2A and 5-HT₇ receptors are heterogeneously distributed in different motoneurons of Onuf's nucleus innervating different striated muscles. 5-HT_2A receptor mainly distributed in motoneurons innervating IC might be preferentially involved in regulation of the sexual reflex, and the 5-HT₇ receptor mainly distributed in motoneurons innervating EUS may be mainly participated in the micturition reflex. Strong GAL immunoreactivity expressed in the medial and lateral parts of the DLN, which involved in innervating the EUS and the IC, may be participated in the urogenital reflex.