• Journal of Acupuncture Research
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• v.21 no.5
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• pp.1-12
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• 2004

Objective : This study was to investigate the effect of acupuncture on NADPH-diaphorase and nNOS in the brain stem and cerebellum of spontaneously hypertensive rats. Methods : The experimental groups were divided into four groups : Normal, Choksamni(ST36), Kokchi(LI11), arbitrary group. Thereafter we evaluated changes in NADPH-diaphorase positive neurons histochemically and changes in nNOS neurons immunohistochemically. Results : 1. The optical densities of NADPH-diaphorase positive neurons of all the Choksamni & Kokchi groups were significantly different in SuG, DLPAG, IP, Pr, Gi areas of brain stem and cerebellum as compared to normal & arbitrary groups. In PPTg only Choksamni group was significantly different as compared to normal and arbitrary groups. 2. The optical densities of nNOS-positive neurons of Choksamni & Kokchi groups were significantly different in SuG, DLPAG areas of brain stem as compared to normal group. In IP, Pr only Kokchi group was significantly different as compared to normal group. The optical densities of nNOS-positive neurons of Choksamni & Kokchi groups were significantly different in SuG, DLPAG, PPTg, Pr, Gi areas of brain stem as compared to arbitrary group. In IP, Pr only Kokchi group was significantly different as compared to arbitrary group. 3. The optical densities of nNOS-positive neurons of all the Choksamni & Kokchi groups were not significantly different in cerebellum as compared to normal & arbitrary groups. Conclusions : We found out that acupuncture have effects on NADPH-diaphorase and nNOS in the brain stem and cerebellum of spontaneously hypertensive rats.

• Biomolecules & Therapeutics
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• v.3 no.1
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• pp.58-62
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• 1995

We investigated whether nitric oxide (NO) may serve a role in bladder function by immunohistochemical analysis of the distribution of intrinsic NADPH-diaphorase and functional study of isometric tension recordings via a photo-induced adequate nitric oxide (PIANO) generating system using rat bladder. Results suggest that a small number of NADPH-diaphorase-positive perikarya are present within the bladder wall and within adjacent small ganglia. Furthermore, NADPH-diaphorase-positive nerve fibers were observed in the adventitial and muscular layers, subjacent to the urothelium and perivascular fibers. Rat bladder strips precontracted with 3$\mu$M carbachol were reversibly relaxed upon NO generation by UV irradiation. PIANO-mediated relaxation was sensitive to oxygen free radicals. In addition, tissue cGMP levels were increased by the PIANO generating system and elevated cGMP levels were decreased by pretreatment of guanylate cyclase inhibitor, methylene blue. These results indicate that NO may serve a role in modulating bladder tone in the rat.

• Journal of Acupuncture Research
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• v.20 no.6
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• pp.148-159
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• 2003

Objective: This study examines the effects of acupoint combination on NADPH-diaphorase and neuronal nitric oxide synthase(nNOS) in the brainstem of spontaneously hypertensive rats. Methods: The changes of NADPH-d-positive neurons using a histochemical method and the changes of nNOS-positive neurons using an immunohistochemical method were evaluated. The optical densities of NADPH-d-positive neurons and nNOS-positive neurons of the Choksamni(ST36) Umnungchon(SP9) groups were significantly increased in all brainstem areas as compared to the Choksamni and Choksamni Kokchi(LI11) groups and decreased, with the exeption of the nNOS-positive neurons in the superficial gray of superior colliculus, as compared to the normal group. Results: Our results demonstrated that electroacupuncture changes the activity in the NO system in the brainstem of SHR and the acupoint combination is one of the important parameters for this effect.

• Journal of Nutrition and Health
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• v.33 no.5
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• pp.491-496
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• 2000

Food intake is regulated by both central and peripheral mechanisms. In the central nervous, the hypothalamus acts for autonomic and endocrine homeostasis. The paraventricular nucleus(PVN) of hypothalamus is an imprtant site of interaction in central feeding pathways. Neuroepetide Y(NPY)is one of the most powerful neurochemical stimulants of food intake known. Also brain nitric oxide(NO), known as neurotransmitter, is involved in the mechanisms that regulate food intake. In this experiment, 24h fasting mice and anorexia mutant mice have been to examine the expression of NPY, which is the major neuropeptide increasing food intake. Double staining with NPY and nicotinamide-adenine-dinucleotide-phosphate diaphorase(NADPH-d), followed by immunohistochemical method and image analysis, have been used to observe coexisting neurons and the level of expression of each neurons. The results were as follows. 1) NPY-immunoreactivitys reduced immune response of the hypothalamus, particularly paraventricular nucleus(PVN), in anorexia mutant mice. Decreased level of NPY is assumed to be a major pathological factor in anorexia mutant mice. On the other hand, PVN in hypothalamus of fasting mice showed increased immunoreactivity which is in agreement of other researchers. 2) NPY and NADPH-d double staining revealed coexisting neurons in the cerebral cortex. Fasting mice had a tendency to have increased level of coexisting neurons compared to the control group. Compared to the control group, fasting mice express is not increase level of NPY-immunoreactivity, while anorexia mutant mice tended to have a decreased level.

• Toxicological Research
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• v.13 no.3
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• pp.251-256
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• 1997

Toluene inhalation increases glutamate level and its receptor in various brain regions. In this study, nitric oxide synthase (NOS) activities were investigated in various rat brain regions using NADPH diaphorase staining method which examined histochemical changes of NOS in the neural cells. Also, in vitro LDH leakage assay and MTT test were performed to investigate the toxic influences of toluene in cultured granule cell of rat cerebellum which was significantly affected with toluene in vivo. Rats were exposed to toluene of 10000 ppm for 3 days. 7 days and 14 days by 20 min $\times$ 2 times a day. NADPH diaphorase staining was processed in the different brain regions after inhalation. NADPH diaphorase staining density was not significantly changed at 3 days inhalation group, but the density decreased in proportion to the duration of toluene inhalation. Over 30% of staining density was decreased at 14 days group which was maximum duration of inhalation in this study. The tendency of staining density decrease was significant in granule cell of cerebellum. Cell death by toluene exposure was observed in cultured cerebellar granule cell. $EC_{50}$ measured with LDH leakage assay and MTT test were 43 mM and 72 mM respectively.

• Journal of Pharmacopuncture
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• v.6 no.2
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• pp.57-65
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• 2003

Objective : This study was carried out to systemically investigate the effect of Joongwan(中脘; CV12) acupuncture in cerebrum and cerebellum of indomethacin-induced gastrointestinal disease in SD-rats. Method : 1. We induced gastrointestinal disease by indomethacin oral administration in SD-rats. 2. We selected Joongwan(中脘; CV12) acupuncture point that generally have been used to treat gastrointestinal disease. 3. We categorized SD-rats into three groups as followings. (1) Normal group : The group without any management (2) Control group : The group with indomethacin-induced gastrointestinal disease (3) Treated group : The group that Joongwan(中脘; CV12) acupuncture was performed after inducing gastrointestinal disease 4. We figured out the effect of acupuncture by analyzing staining degree of NADPH-diaphorase in cerebrum and cerebellum. Results : 1. Cerebrum (1) Normal group : The degree of staining was very low. (2) Control group : NADPH-diaphorase was mainly stained in cerebral cortex and the stained region was wider than Normal group. (3) Treated group : The degree and region of staining was higher and wider than the other goups. Sometimes the intensively stained regions were observed. 2. Cerebellum In both cases of Control group and Treated group, the regions in cortex were stained mainly. But, between Control group and Treated group, there was no remarkable difference. Conclusion : In case of cerebellum, there was no remarkable result. On the other hand, in case of cerebrum, there were certain differences among three groups. Through those results, we could conclude that Joongwan(中脘; CV12) acupuncture treatment was able to affect NADPH-diaphorase expression in the cerebrum of SD-rats that have gastrointestinal disease with indomethacin-inducing.

• Korean Journal of Biological Psychiatry
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• v.3 no.2
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• pp.191-202
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• 1996

This study was aimed to charily the change of NADPH-diaphorase(NADPH-d) and neuropeptide Y(NPY) associated with aging of the rots. To verily the effect of aging of NPY and NADPH-d-positive neurons in the brain stem, the neurons were stained by the immunohistochemical and histochemical method. In the aged group, the number of NADPH-d-positive neurons was significantly decreased in substantia nigra lateralis, supragenual nucleus, raphe magnus nucleus and raphe obscurus nucleus as compared with control group. The number of NADPH-d-positive neurons was not significantly decreased in superior colliculus, interpeduncular nucleus, central gray. dorsal raphe nucleus, retrorubral nucleus, pedunculopontine tegmental nucleus, laterodorsal tegmental nucleus, pontine reticular nucleus, prepositus hypoglossal nucleus and nucleus solitarius of the aged rats. The NADPH-d and NPY-positive neurons were found in the interpeduncular nucleus, central gray, substantia nigra lateralis, laterodorsal tegmental nucleus, nucleus solitarius, raphe magnus nucleus, raphe obscurus nucleus of the control and aged groups. The coexistence of NADPH-d and NPY in the some cell was not found in the brain stem of both groups.

• Applied Microscopy
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• v.45 no.3
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• pp.135-143
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• 2015

The aim of this study was to investigate effects on joint mobilization in neurochemical changes of nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) and nitric oxide synthase (NOS) of the spinal cord neurons after right knee joint immobilization (RKJI) and in ultrastructural changes of the femoral nerves innervating the muscles acting on RKJI. A total of 15 guinea pigs were used and divided into 5 groups. Immunohistochemistry was performed to detect NADPH-d and NOS. NADPH-d and NOS were not expressed in the ventral horn of control and experimental groups, but were expressed or not in the dorsal horn according to the duration of release after RKJI and the presence or absence of joint mobilization. Ultrastructures of the femoral nerves in experimental groups had partial demyelination and condensed clumps in axon. Effects on manipulative therapy after RKJI were confirmed from expression of NADPH-d and NOS in the dorsal horn of the lumbosacral spinal cord. Manipulative therapy was more effective against a long-term immobilization than a short-term immobilization.

• Journal of Acupuncture Research
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• v.25 no.3
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• pp.179-187
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• 2008

Objectives & Methods : This study was to investigate effect of low frequency electroacupuncture on NADPH-d positive neurons in the brain cortex of rat with adjuvant induced rheumatoid arthritis. Experimental groups were divided into 6 groups ; Normal, Control, $ST_{36}$, $SP_9$, $ST_{36}+SP_9$ and Non-Acupoint. Normal group, non-arthritic group, was injected normal saline, and the other groups were injected FCA. Each acupoint groups were treated by 2Hz electroacupuncture at each acupoints and NA group was treated by 2Hz electroacupuncture at non-acupoint. Each groups were evaluated by the number of NADPH-d positive neurons in primary somatosensory area(S1), secondary somatosensory area(S2), motor area and caudate putamen by using an image analyzer and a microscope. Results : 1. In S1, the number of NADPH-d positive neuron cells in the $ST_{36}$ group were significantly(p<0.05) increased compared with the control group. 2. In S2, the number of NADPH-d positive neuron cells in all electroacupuncture groups were not significantly changed compared with the control group. 3. In motor area, the number of NADPH-d positive neuron cells in $ST_{36}$ group, $SP_9$ group, NA group were significantly(p<0.05) increased compared with the control group. 4. In Caudate putamen, the number NADPH-d positive neuron cells in all electroacupuncture groups were significantly(p<0.05) decreased compared with the control group. Conclusions : Our result demonstrated that low frequency electroacupuncture on $ST_{36}$ & $SP_9$ normalized expression of NADPH-d positive neurons in the brain cortex of the rheumatoid arthritis model in rats.

• Journal of Acupuncture Research
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• v.18 no.2
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• pp.79-90
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• 2001

Objective : The aim of the present study is to investigate whether stimulating a specific auricular point(胃點) is effect on suppression of appetite. Methods : This study evaluated the changes of Reduced-Nicotinamide-Adenine-inucieotide-Phosphate-diaphorase(NADPH-d)-positive neurons using a histoehemical method. Staining intensities of NADPH-d-positive neurons were assessed in a quantitative fashion using a microdensitometrical method based on optical density by means of an image analyzer. Results : The results show as follows ; 1. In the RSG/RSA, FRI/FR2, HL, PAR1 area, The optical density of NADPH-d-positive neurons of fasted group was significantly decreased in comparison to fed group. 2. In most cortical area, It was not significant statistically, But the optical density of NADPH-d-positive neurons of fed with auricular acupuncture group was decreased in comparison to fed group. 3. In most cortical area, It was not significant statistically, But fasted with auricular acupuncture group increased the optical density of NADPH-d-positive neurons comparison to fasted group. Conclusions : We conclude that stimulating the specific auricular points is able to effect on NO(Nitric Oxide)-expression in the brain cortex of sprague dawley rats, The results suggest that auricular acupuncture may be useful for controlling obesity.