Objective To investigate the possible role of orexin in reproduction and examine the changes in the expression of orexin and its receptors(OX1R,OX2R) in the rat hypothalamus during pregnancy,parturition and lactation. Methods The expressions of prepro-orexin(prepro-OX),orexin-A,OX1R,and OX2R in the rat hypothalamus during pregnancy,parturition,and lactation were evaluated by competitive reverse transcription-polymerase chain reaction(RT-PCR) and immunohistochemistry assay. Results Orexin-A immunoreactive(ir) neurons and the OX1R subtype in neuronal cell bodies were mainly located in the lateral hypothalamic area(LHA) as well as in the magnocellular neurons of the paraventricular and supraoptic nuclei respectively in pregnant and lactating rats.The prepro-OX and OX1R mRNA levels on the 1st day of lactation were significantly higher than that during late pregnancy and lactation.No significant changes of OX2R expression were observed during the various reproductive phases.Conclusion Orexin might be involved in regulating reproductive function in early lactation through their binding sites in hypothalamic PVN and SON.

BACKGROUND: Quite a few deoxidized reduced form of nicotinamide-adenime dinucleotide phosphate (NADPH-d) -positive neurons distribute in amygdala complex (AMC) and gamma-aminobutyric acid(GABA)is an important inhibitory neurotransmitter that also widely distributes in the central nervous system (CNS) of mammals. Whether there is coexistence of GABA and nitricoxide synthase (NOS) in AMC is unknown at present.OBJECTIVE: To observe whether there is GABA and NADPH-d-eoexist neuron in cortical nucleus of amygdala (Co) with the combination of NADPH-d hischemical and immunohistochemical double staining.DESIGN: A verifying controlled study based on the experimental animals.SETTING: Department of anatomy of two universities MATERIALS: The study was conducted in the department of Anatomy,Medical College of Zhejiang University between May 2004 and June 2004. Six SD rats in either gender with a body mass between 250 g and 300 g were selected.INTERVENTIONS: Coronary serial frozen slices of brain tissue were prepared. 4 sets of slices were selected for Nissl staining, NADPH-d histochemical staining, NADPH-d histochemical and GABA immunohistochemical double staining, and control experiment for the test of antibody specificity. GABA-labeled neurons, NADPH-d-positive neurons, and NADPH-d/GABA double-labeled neurons were counted in Co nucleus for the calculation of the percentage of double-labeled positive neurons to single-labeled positive neurons.MAIN OUTCOME MEASURES: Distribution of NADPH-d/GABA double labeled neuron; distribution of NADPH-d or GABA single labeled neuron.RESULTS: Most of GABA-positive neurons distributed in posteromed cortical amygdaloid nucleus (PMCo), posterolateral cortical amygdaloid nucleus (PLCo), which were mainly small types but few middle types. Most of NADPH-d-positive neurons distributed in PMCo, PLCo and anterior cortical amygdaloid nucleus (Aco), which were mainly middle sized and small sized neurons. The ratio of NADPH-d/GABA double-labeled neurons to single labeled neuron was 24.5% (GABA) or 46.7% (NADPH-d).CONCLUSION: There is GABA/NADPH-d-coexist neuronsin Co of rats,which suggests NO might have modulative effect on GABA neuron in Co.

Objective To estimate the effect of the NO on the medial amygdaloid nucleus(Me), we studied the origins of the NOS positive terminals in the Me. Methods Noergic afferent projection to Me was identified by a combined NADPH-d histochemical staning and retrograde CTb immunocytochemical method after microinjecting CTb into Me. Results The double labeled of neurons (NOS and CTb) were located in dorsal raphe nucleus, locus ceruleus, basolateral amygdaloid nucleus, parabrachial nucleus, ventrolateral part of periaqueductal gray.Conclution The NADPH-d positive terminals in the Me originates from the aforementioned nucleus, and may relate to the function of the Me.

Objective In the present study, we observed the distribution of nitric oxide synthase positive neurons in the developing rat telencephalon and to realize the possible roles of NO in brain development further. Methods NADPH-d histochemical method was used to investigate the micromorphology and distribution of NOS positive neurons in rat telencephalon from embryonic day 14 (El4) to postnatal day 14 (P14). Results At El4, no neurons stained. Labeling was observed in the ventrolateral of the striatum at E15. At E17, positive neurons were seen in the cerebral cortex and piriform cortex. They were cells with small cell bodies, shorter dendrites and less ramification. The stained neurons were either increased or maintained in both number of neurons and intensity during development. At E20, positive cells were found in many regions. Many positive neurons were present in the piriform cortex, ventrolateral of the striatum and bed nucleus of the stria terminalis. They acquired larger size and more complex dendrites. After birth, intensely stained NOS positive fibers were distributed widely in the cerebral cortex and striatum. Stained cells were found in the corpus callosum and hippocampus also. The distribution pattern of positive cells at P14 was essentially similar to that reported in adult rats. Conclusion The distinctive temporal and spatial expression pattern of nitric oxide synthase positive neurons in the telencephalon indicated that NO may play an important role in brain development and maturation.

Objective To investigate a possible role of orexin in the regulation of estrous cycle by examining the expression of orexin and orexin receptors(OX1R,OX2R) in the rat hypothalamus during the estrous cycle. Methods The levels of prepro-orexin(prepro-OX),orexin-A,OX1R,and OX2R in the rat hypothalamus during the estrous cycle were evaluated by competitive reverse transcription-polymerase china reaction(RT-PCR) assay. Results Only expression OX1R mRNA during late proestrus was significantly higher than that at metestrus.No significant changes of prepro-OX and OX2R mRNA expression were observed during the various estrous cycle phases.Conclusion Orexin might regulate the secretion of GnRH and/or LH by binding OX1R,contributing to the occurrence of ovulation.

Objective To investigate the expression of regeneration gene protein-2(Reg-2) after the transection injury in rat spinal cord. Methods Forty five SD rats were randomly divided into two groups: in the injury group,the SCI was produced by complete transection of the cord at the 9(th) thoracic level,and the sham operation rats were taken as the normal control.The animals were perfused at 1h,1d,3d,7d,14d after the operation,and the spinal cord was taken out at different time points.The expressions of Reg-2 were tested by immunohistochemical analysis and Western blotting,the double staining of Reg-2 and different neural cells special markers(GFAP,Olig2,NPY,CGRP,GAP-43,et al)was done with immunofluorescence method. Results The expression of Reg-2 was found at 1h after the spinal cord injury,reached the peak on day 3 after the injury in the neurons of dorsal horn and day 7 in the neurons of ventral horn.The high expression persisted for 1 week,then decreased gradually.Conclusion Reg-2 may work as an important growth factor and participate in the regeneration and rehabilitation process after the spinal cord injury.

Objective To observe the distribtuion of NOS positive neurons in the projecting pathway to the subnucleus reticularis dorsalis(SRD) in the rat. Methods We used a double staining technique combined retrograde tract tracer and NADPH d histochemistry. Results FG/NOS double labeled neurons were found in the laminae Ⅰ,Ⅹ of spinal cord,dorsal raphe nucleus(DR),linear raphe nucleus(LR),and raphe obscurus nucleus(RO).In the periaqueductal gray(PAG),there were many FG labeled or NOS labeled neurons,but no FG/NOS double labeled neurons were found.Conclusion\ NO might play a role in the pathway of afferent projections of the SRD.\;[

Objective To investigate the distribution of moderate to severe cancer pain and administrated dose of opioid drugs in order to provide the reference for the treatment of moderate and severe cancer pain.Methods Retrospective analyzed the clinical data of 146 patients with moderate to severe cancer pain.The information were recorded including opioid use dose,cancer pain incidence,age,gender,height,weight,body surface area and type of tumor.Results There were no significant difference among patients with opioids drug administration in terms of gender,average daily oral dose(x2 =0.473,Z =-0.185,P ＞ 0.05).Meanwhile cancer incidence rate was same in different age groups (x2 =2.280,P =0.684).The average daily opioid dose in patients with over 80 year old was the lowest among other age groups (Z =-2.745,-1.986,-2.141,-2.162;P ＜0.05).Cancer pain incidence in patients with hepatobiliary pancreatic tumors were 67.50％ (27/40),highest than other different types cancer including metastatic tumor(62.50％ (10/16)),urinary tumor (61.54％ (8/13)),lung cancer (52.63％ (50/95)),breast cancer and gynecological tumor (50.00％ (17/34)),gastrointestinal tumor(41.94％ (26/62)),other tumors 38.46％ (5/13) and head and neck tumor(25.00％ (3/12)),the difference was statistically significant (x2 =23.672,P ＜ 0.05).The average day oral morphine dose of different types tumor were listed as followed from high to low in the order:160 (80,200) mg of metastatic tumors,120 (60,160) mg of breast and gynecological tumor,100 (40,125) mg of lung cancer,90(45,115) mg of urinary tract tumors,80(60,160) mg of other tumors,70(50,90) mg of hepatobiliary pancreatic tumor,60 (40,80) mg of gastrointestinal tumor,55 (40,70) mg of head and neck tumor.There were significant differences in terms of among oral morphine dose per day of different type tumors (H=14.280,P ＜0.05).No correlation was found between pain patients with height,body mass,body surface area and average daily dose of morphine (r =-0.045,-0.042,-0.046 ; P ＜ 0.05).Conclusion No significant differences were found in terms of moderate to severe cancer pain among different type tumors at different and age groups.While there is significant difference in term of cancer pain incidence among different tumor types.The average daily amount of morphine in patients with cancer pain is not related to gender,height,body mass and body surface area.The average amount of morphine of patients with age over 80 years old is significantly lower than that of the other age groups,and the average amount of morphine t is related to cancer type.

Objective To study the expression of substance P(SP) and substance P receptor(SPR) during the development of mice brains. Methods The expression of SP and SPR during the development of mice brains from embryonic day(E) 11 to postnatal day(P) 0 days was analyzed by immunohistochemical method. Results The expression of SP began at E11 and gradually increased until birth. The expression of SPR began at E11 and maintained stable expression until birth. SP mostly expressed at striatum and SPR mostly expressed at medullary raphe.Conclusion The expression of SP and SPR during the embryo brain stage may indicate that SP could be an important factor involved in the early organization and maturation of neuron.

BACKGROUND: Frenulum of prepuce of penis contained many nerve terminals is an extremely sensitive region. If the frenulum is injured in circumcision or other operations, the complication, such as postoperative spontaneous pain of penis, sexual disturbance and so on, will occur. But there still is no define explanation for this up to now.OBJECTIVE: To observe the distribution of immunoreactive nerve terminal of calcitonin gene-related peptide (CGRP) in prepuce of penis and frenulum of prepuce of adult SD rats, and look for the source of CGRP immunoreactive nerve terminal in frenulum of prepuce.DESIGN: A single sample trial.SETTING:Department of Anatomy, School of Medicine, Zhejiang University.MATERIALS: The experiment was performed at Department of Anatomy,School of Medicine. Zhejiang University from September 2004 to May 2005. A total of 20 adult male SD rats were selected, and were raised in warm, quiet, photophygous environment for 1 week before the trial so as to make the rats fit for the environment and maintain their basal state.METHODS: The rats were assigned randomly into 2 groups. Ten rats in the first group were treated with the immunohistochemical method to observe the distribution of CGRP immunoreactive nerve terminal in prepuce of penis and frenulum of prepuce of adult rats. Ten rats in the second group were treated with fluorogold (FG) retrograde labeled combined with CGRP immunofluorescence labeled method to look for the source of CGRP immunoreactive nerve terminal in frenulum of prepuce of penis. MAIN OUTCOME MEASURES: ①The morphology and distribution of CGRP immunoreactive nerve terminal in prepuee of penis and frenulum of prepuce of adult SD rats were observed under light microscope. ②The distributive density and difference of CGRP immunoreactive nerve terminal in prepuce of penis and frenulum of prepuce were detected and compared (represented by A). ③Morphology and distribution of FG retrograde labeled -positive, CGRP single-labeled positive and FG/CGRP double-labeled positive neurons in dorsal root ganglion were observe under fluorescence microscope. ④Mean quantity of FG retrograde labeled positive, CGRP single abeled positive and FG/CGRP double-labeled positive neurons in dorsal root ganglion was counted.RESULTS: Totally 20 rats were involved in the analysis of results. ① Amber-coloured CGRP immunoreactive nerve terminal appeared in prepuce of penis and frenulum of prepuce of adult rats. These nerve terminal mainly occurred in basal layer of epidermis and papillary layer of dermis, distributed as twig shape or intestiniform; mostly of them were bundled, different in length, and some of them showed enlarged nodosity. ②The distributive density of CGRP immunoreactive nerve terminal in frenulum of prepuce of penis was significantly larger than that in prepuce of penis (2.15±0.32, 1.02±0.22,t =-2.03,P＜0.01). ③Combined with the FG retrograde labeled method it was found that these nerve terminal was derived from neurons of dorsal root ganglion opposed to the sixth lumbar spinal cord and the neurons of dorsal root ganglion opposed to the first acral spinal cord. FG retrograde labeled positive neurons differed in length. The cell body showed round or orbicular-ovate, without obvious prominence. Bright inaurate fine particle appeared in cytoplasm, no label in nucleus. Most cells arranged in line along nerve tract or diffusedly distributed. Most CGRP single-labeled positive neurons were middle or small cells found by CGRP immunofluorescence labeling. Dyeing was too dark.Reaction product distributed evenly in cytoplasm, which showed bright dark green (FITC labeled color). The same positive section was observed comparatively under different excitation light. It was found that FG/CGRP double-labeled positive cells were middle or small, and its amount accounted for a half of the total number of FG retrograde positive cells.CONCLUSION: CGRP may participate the transmission of sensory information in prepuce of penis and frenulum of prepuce of rats. The CGRP immunoreactive nerve terminal in frenulum of prepuce of penis of rats is sourced from neurons of dorsal root ganglion opposed to the sixth lumbar spinal cord and the neurons of dorsal root ganglion opposed to the first sacral spinal cord.