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Objective To study the distribution characteristics and relationship of acetylcholinesterase (AChE) .substance P(SP), vasoactive intestinal polypeptide (VIP) and nitric oxide synthase ( NOS)-positive neurons in intestinal tract of rats. Methods The distributions of AChE,SP, VIP and NOS-positive neurons in duodenum,jejunum,ileum,colon and rectum of rats were respectively observed by using enzyme histochemical and immunohistochemical methods. Results Four types of positive neurons distributed throughout in each laminae of intestinal tract of rats,and there was accompanying and resemble distribution regularity. From duodenum,jejunum to ileum,four types of positive neurons showed gradual increase tendency in quantity, while from colon to rectum, they showed gradual decrease tendency in quantity. There were extremely abundant AChE-posi-tive neurons in each laminae of intestinal tract of rats,comparatively rich NOS and SP-positive neurons in Auerbach'myenteric plexus(AMP) ,and comparatively rich VIP-ir positive neurons in the Meissner's submucosal plexus(MSP)and mucosa. Density on the distribution of four types of positive neurons was AChE>NOS>VIP>SP. There was coexistence of four types of neurons, and there were synapse-like contacts in four types of neurons in AMP. Conclusion There were significant differences in the density on the distribution of AChE,SP, VIP and NOS-positive neurons in different segments of intestinal tract of rats,and there was close morphological relationship among four types of neurons in AMP. It was deduced that they exerted synergistic effects in regulating motility of the intestine.
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In order to investigate the protective effect of hypoxic preconditioning on the cerebral ischemia-reperfusion injury, the expression of Bcl-2 and Bax was detected by using immunohistochemical staining after 3 h cerebral ischemia followed by 1, 6, 12, 24 and 48 h reperfusion respectively in rats treated with or without hypoxic preconditioning before cerebral ischemia. In addition,the apoptosis of neural cells and the behavioral scores for neurological functions recovery were evaluated by TUNEL staining and "crawvling method", respectively. Compared with control group (cerebral ischemia-reperfusion without hypoxic preconditioning), the expression of Bcl-2 was significantly increased, but that of Bax decreased in the hypoxic preconditioning group (cerebral ischemiareperfusion with hypoxic preconditioning), both P＜0. 05. The pre-treatment with hypoxic preconditioning could reduce the apoptosis of neural cells and promote the neurological function recovery as compared to control group. It was suggested that hypoxic preconditioning may have protective effects on the cerebral ischemia-reperfusion injury by inhibiting the apoptosis of neural cells, increase the expression of Bcl-2 and decrease the expression of Bax.
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Objective: To investigate the mechanism of improvement effect of electro-acupuncture at acupoint Neiguan (PC 6) on acute myocardial ischemia (AMI). Methods: Ninety-six rats were randomly divided into sham-operation group, myocardial ischemia model group and myocardial ischemia model plus electro-acupuncture group, the rat model of AMI was established by occlusion of the anterior descending branch of the left coronary artery, then electro-acupuncture therapy was performed at bilateral acupoint Neiguan (PC 6). The myocardial enzyme activities in plasma was detected with biochemical method, and the myocardial c-fos gene expression was detected with reverse transcription polymerase chain reaction(RT-PCR) method. Results: The myocardial enzyme activities in plasma and the myocardial c-fos gene expression in myocardial ischemia model group were obviously higher than those in sham group(P＜ 0.05), after electro-acupuncture treatment, the myocardial enzyme activities in plasma and the myocardial c-fos gene expression were decreased,there was a significant difference (P＜ 0.05). Conclusion: The mechanism of improvement effect of electro-acupuncture at acupoint Neiguan (PC 6) on AMI is related to down-regulation of the myocardial c-fos mRNA expression.
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BACKGROUND: In China, this laboratory is the first one to report such researches, confirming that strong αo-immunoreactive (IR) appears in the substantia gelatinosa (SG) of spinal cord and lateral spinal nucleus which is similar to the distribution of certain neuropeptides that participate in sensory regulation, which suggests that guanine nucleotide binding protein (G protein) may be related to primary afferent informational transfer. OBJECTIVE: To observe the change of αo-IR in gelatinous substance by the method of transection of unilateral spinal dorsal roots.DESIGN: A randomized controlled experiment on animals.SETTING: Staff Room of Neurobiology, Tongji Medical College, Huazhong University of Science and Technology.MATERIALS: The experiment was conducted at the Staff Room of Neurobiology, Tongji Medical College, Huazhong University of Science and Technology from December 1995 to December 1996. Fifteen healthy adult SD rats were selected and divided into 3 groups: ①normal group with five rats (not dealt with any disposal), ②transected dorsal root group with 10 rats (right side) and ③control group (non-transected left sidedness as control).METHODS: Right lumbar 1-3 spinal neural dorsal roots were cut off under the anesthesia of 100 g/L chloral hydrate (300 mg/kg)through intraperitoneal injection in rats, living for 48-60 hours after operation. The subunit αo of guanine nucleotide-binding protein (rabbit polyclonal antiserum) was demonstrated in the αo-IR of rat spinal cord by immunohisto chemical methods. G protein was oriented, and its change was observed after transection ofneural dorsal roots MAIN OUTCOME MEASURES: ①The αo-IR of Ⅰ to Ⅲ of the dorsal horn and lateral spinal nucleus of the normal rats and control rats. ②The αo-IR of Ⅰ to Ⅲ of the dorsal horn and lateral spinal nucleus of rats in the transected dorsal root group. RESULTS: Data of a total of 15 rats were involved in the result analysis. ①In the normal group and control group, intense αo-IR was presented in rexed lamina ( Ⅰ to Ⅲ ) of the dorsal horn of rats, and the highest αo-IR in second lamina (SG). Lateral spinal nucleus of rat revealed higher density of αo-IR containing fiber networks. Following unilateral transection of dorsal roots in SG, αo-IR was markedly decreased. ②Quantitative analysis of absorbance (A) of αo-IR, it was (0.847±0.081) in the inside of the control group, (0.633±0.073)(t=5.71 ,P ＜ 0.001 ) in the inside of transected dorsal root group. It was (0.823±0.089) in the middle area of the control group,(0.660 4±0.074)(t=6.90,P ＜ 0.001 ) in the middle area of the transected dorsal root group. It was (0.915±0.090) in the lumbar region of the control group, and (0.656±0.077)(t=10.31 ,P ＜ 0.001 ) in the lumbar region of the transected dorsal root group. Average value of the control group was (0.852±0.084), and average value of the transected dorsal root group was (0.639±0.078)(t=10.23 ,P ＜ 0.001 ).CONCLUSION: Part of G protein of end-brush neurons related with the primary afferent noxious stimulation in SG derives from primary sensory neurons, which maybe join the adjustment of primary sensory transfer.
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BACKGROUND: The effect of external application of capsaicin in the treatment of superficial pain has been recognized, but its effect against trigeminal neuralgia by direct action on the nerve ending or nerves in the hypodermis or deep tissues awaits intensive investigation.OBJECTIVE: To observes the effect of subcutaneous injection of capsaicin on calcitonin gene-related peptide (CGRP) and nitric oxide synthase(NOS)-positive nerve fibers in rat facial skin.SETTING: Teaching and Research Division of Anatomy, Xianning Medical College, and Department of Neurobiology, Tongji Medical College of Huazhong University of Science and Technology.MATERIALS: This experiment was conducted in the Laboratory of Neurobiology, Tongji Medical College, between October and December 2003.Twenty healthy Wistar rats of either sex with body mass of 120-170 g were used.METHODS: The rats received subcutaneous injection of capsaicin for treatment of the suborbital branch of the trigeminal nerve on the right side,with the left side serving as the control side. According to the doses of capsaicin applied, the rats were divided into 4 groups, namely 20, 30, 50and 100 μL capsaicin groups with 5 rats in each group. Twenty-four hours after the injections, samples were obtained and cut into slices for microscopic observation and the expressions of CGRP and NOS were examined immunohistochemically.MAIN OUTCOME MEASURES: [1] Changes of CGRP and NOS-positive nerve fibers on the experimental side and image analysis of the mean absorbance of CGRP and NOS; [2] changes of characteristic behaviors and body signs of the rats.RESULTS: Totally 20 rats entered the result analysis. [1] Behavioral change: A few minutes after subcutaneous injection of capsaicin, the rats exhibited a series of characteristic behavioral and symptomatic changes,which gradually diminished or even vanished with the increase of the doses. [2] Microscopic changes: On the experimental side, no obvious difference was noted in the expression of CGRP and NOS-positive nerve fibers between the groups. [3] Imaging analysis of the mean absorbance of CGRP and NOS: For CGRP, the mean absorbance was 0.984±0.056 on the control side and 0.947±0.025, 0.852±0.042, 0.756±0.028 and 0.730±0.016 in 20,30, 50 μL and 100 μL capsaicin groups, respectively. As for NOS, the mean absorbance was 0.151±0.009 on the control side, and was 0.148±0.007,0.132±0.012, 0.111±0.067 and 0.107±0.006 in 20, 30, 50 μL and 100 μL capsaicin groups. Analysis of variance revealed significant differences between the groups (P ＜ 0.01).CONCLUSION: CGRP and NOS participate in the processing of nociceptive information and modulate pain and analgesia. Capsaicin executes analgesic effect by exhausting massive neurotransmitters.
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Objective:The stimulative effects of clonidine on release of enkephalin and substance P from periaqueductal gray was studied to analyse the analgesic neurotransmitters related to alpha 2 adrenoceptor activation. Method: Twenty SD rats were allocated in pairs to receiving intraperitoneal clonidine 40?g in clonidine group or normal saline in control group. Immunohistochemical technique was used to measure the contents of enkephalin and substance P in slice of rat midbrain. Result:The immunoreaction of enkephalin from slice of rat midbrain was decreased significantly in clonidine group compared with control group(P
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The subunit ?_0 of guanine nucleotide- binding protein, in the areas of rat brain and spinal cord was localized by immunohistochemical methods. It was found that in the rat brain, specific ?_0-like immunoreactivity(?_0-Li) displayed regional heterogeneity, a high density of ?_0-Li revealed in neuropil, and somatic membranes as well as the neuronal processes.Most intense ?_0-Li can be seen in substantia nigra(pars reticulata), interpcduncular nucleus, habenulo-interpeduncular tract, strata oriens and radiatum of the hippocampus, and substantia gelatinosa of the spinal cord. There are also areas of moderate staining ie: the molecular layer of cerebral and cerebellar cortex, habenula, caudate-putamen complexes, the midline nuclei of thalamus and hypothalamus, periaqueductal grey, grey layers of superior colliculus, the olivo-cerebellar tract and the spinal tract of the trigeminal nerve as well. By contrast, the immunoreactvity of ?_0 in septal nuclei, globus pallidus, red nucleus, and the regions adjacent canalis centralis of the spinal cord showed much weaker. In addition, on the membranes and the processes of the neuronal cell bodies in the periaqueductal grey, substantia nigra, reticular formation, medial geniculate body and the nucleus of the trapezoid body were ?_0-Li positive.The results of AChE staining revealed that the AChE-positivc nerve terminals was coinsident with the presence of ?_0-Li in the following regions. For instance: the molecular layer of cerebral cortex, hippocampus, spinal tract of the trigminal nerve, and substantia gelatinosa of the spinal cord, where both the ?_0-Li and the AChE activity were positive. It is suggested that ?_0 subunit of Go-protein in brain might play roles in membrane signal transduction, and might have some relationship with cholinergic nerve.
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In this study histochemical method was used. It has been found that on the seventh day after epidural administration of capsaicin substance p like immuno- reactivity (SPAR) in laminae Ⅰ-Ⅱ of dorsal horn of spinal cord decreased obv- iously as compared with that of vehicle treated animals, meanwhile, the activity of fluoride resistant acid phosphatase (FRAP) in both laminae Ⅱ of dorsal horn of spinal cord and the B-type cells of spinal ganglion disappeared. On the cont- rary the Leu-enkephalin immunoreactivity (ENK-IR) in laminae Ⅰ-Ⅲ of dorsal horn of spinal cord apparently increased as compared with that in vehicle group. the evidence from functional measurement showed that capsaicin can raise pain threshold, however, vehicle didnot have obvious influence on pain threshold. The foregoing results suggest that epidural administration of capsaicin in adult rat can effect the level of neurontransmitters, the activity of enzymes and block the con- duction of the primary afferent C fibers; the capsaicin-sensitive C fibers may have relationship with the ENK-containing interneurons in dorsal horn of spinal cord.
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Immunoreactivity of neurotensin(NT) and substance P (SP) were studied by means of PAP method with simultaneous immunocytochemical double staining on the same sections using diaminobenzidine (DAB) for SP and benzidinc dihydrochloride(BDHC) for NT as the chromogens for light and electron microscopy in dorsal horn of spinal cord of rat treated with colchicinc. In our hands the reaction products of DAB and BDHC is quite discernible, under LM the former is brown and later is blue; under EM DAB reaction product is homogeneous and diffuse electron opaque while BDHC reaction product is patch electron opaque, it is a new but simpler approach to demonstrate two antigens simultaneously in the same ultrathin section. It would be conformable further to the studies of morphological and functional relationships between different kinds of neurons. The results showed that NT-like immunoreactive (NT-LI) perikarya were mainly located in lamina Ⅱi and out layer of lamina Ⅲ and NT-LI terminals were mainly located in laminae Ⅰ-Ⅲ. Under EM the NT-LI axon terminals may synapse with the unlabeled axon or unlabeled dendrites. SP-like immunoreactive (SP-LI) perikarya were mainly located in lamina Ⅱ. The density of SP-LI terminals were higher in laminae Ⅰ-Ⅱ, while axo (SP)-axonic (SP), axo (SP)-somatic (SP) synapses were identified with EM. In double labelling sections, furthermore under EM, it was found that NT-LI axon terminal (BDHC patch like electron opaque reaction products)can synapse or contact with SP-LI axon (DAB diffuse electron opaque reaction products). Our results suggested that NT-and SP-containing neurons and terminals in dorsal horn might participate in regulating process of primary sensory transmission.