INCREASED NEURONAL NITRIC OXIDE SYNTHASE EXPRESSION IN THE GRACILE NUCLEUS OF BRAINSTEM FOLLOWING ELECTROACUPUNCTURE GIVEN BETWEEN CUTANEOUS HINDLIMB ACUPUNCTURE POINTS BL 64 & BL 65 IN RATS

This study was to examine the influence of electroacupuncture (EA) stimulation on neuronal nitric oxide synthase (nNOS) expression in the brainstem nuclei in rats. Low-frequency EA stimulation (3 pulses/sec) was applied between 2 acupuncture points (acupoints), Jinggu (BL 64) and Shugu (BL 65) which are cutaneously located at hindlimb, in rats anesthetized with ketamine. After 2 hours stimulation and sham-treatment, animals were perfused with 4% paraformaldehyde. Sections of rat medulla were examined by immunolabeling with a polyclonal antibody directed against nNOS. The brainstem nuclei were also visualized by NADPH-diaphorase histochemistry, a marker of nNOS activity. nNOS levels were quantified by using a microscope with reticule grid to count the number of positive cells over an area. Unilateral EA stimulation of BL 64 and BL 65 in rats caused increases in nNOS immunostaining cells in the ipsilateral and contralateral gracile nucleus compared with sham-treated rats (P<0.05, n=6). NADPH-diaphorase positive cells were also increased in the gracile nucleus of the rats with EA stimulation. Neither nNOS immunostaining nor NADPH-diaphorase reactivity was altered in the nucleus tractus solitarius, rostral ventral medull and other brainstem nuclei in rats with EA stimulation. These results show that nNOS immunoreactivity and NADPH-diaphorase reactivity are consistently increased in the gracile nucleus by low-frequency EA applied to BL 64 and BL 65. We conclude that EA stimulation of the cutaneous hindlimb acupoints induces nNOS expression in the gracile nucleus, and enhanced nNOS-NO in the area may mediate somatosympathetic reflex activities, which contribute to therapeutic effects of acupuncture.