Morphological characteristics of the lumbosacral spinal cord urogenital center in Sprague Dawley rats

BACKGROUND： Studies have demonstrated that selective innervation of the sacral nerve tract to the bladder plays an important role in bladder functional reconstruction following spinal cord injury. However, there are very few studies reporting detailed morphological characteristics of urogenital center and lumbosacral nerve roots. OBJECTIVE： To analyze the spinal cord segment of the lumbosacral spinal cord urogenital center, and to observe morphological characteristics. DESIGN, TIME AND SETTING： A neuroanatomical study was performed at the Laboratory of Neuroanatomy, Peking University Health Science Center between September 2007 and March 2008. MATERIALS： Horseradish peroxidase-conjugated cholera toxin B subunit （CB-HRP） was purchased from Sigma, USA; surgical microscope was purchased from Zhenjian Zhongtian Optical Instrument, Jiangsu Province, China; BCL-420 biological and functional experimental system was purchased from Taimeng Science and Technology, Sichuan Province, China. METHODS： A total of 36 adult, Sprague Dawley rats were randomly assigned to groups A （n = 10）, B （n = 10）, C （n = 10）, and D （n = 6）. CB-HRP （3%, 10-15 μL） was injected into the bladder detrusor muscle （group A）, external urethral sphincter （group B）, and perineal muscles （group C）, respectively. Rats in group D were not given any treatments. MAIN OUTCOME MEASURES： At 72 hours after CB-HRP injection, CB-HRP-positive neurons were analyzed in lumbosacral segments using 3, 3＇, 5, 5＇-tetramethylbenzidine staining and an Olympus optic microscope, while anatomical structures in the respective spinal nerve tract were observed using a surgical microscope. RESULTS： CB-HRP-positive neurons were distributed in the L6-S1 segments of the spinal cord, and neurons primarily innervating the bladder detrusor muscle were located at the sacral parasympathetic nucleus and the intermediolateral nucleus. In addition, neurons that primarily innervate the external urethral sphincter and perineal muscles were observed in the ventrolateral portion （Onuf＇s nucleus）. The lumbar-sacral nerve roots were composed of varying nerve tracts, Le., they were typically divided into 1-2 sub-bundles, and the sub-bundles were then divided into 2-3 tiny bundles. There were extensive fibro-connections between the rootlets. CONCLUSION： The urogenital center in Sprague Dawley rats was located in the L6 -S1 segments of the spinal cord, and the rootlets were clearly observed. Therefore, this rat experimental model could be utilized for highly selective anterior/posterior rhizotomy.

Combined Subchronic Toxicity of Bisphenol A and Dibutyl Phthalate on Male Rats

Objective To evaluate the combined subchronic toxicity of bisphenol A（BPA） and dibutyl phthalate（DBP） in male Sprague Dawley（SD） rats.Methods Forty 4‐week‐old male rats weighing 115‐125 g were randomly divided into BPA‐treated,DBP‐treated group,BPA＋DBP‐treated and control groups and fed with a soy‐ and alfalfa‐free diet containing 285.4 ppm BPA,285.4 ppm DBP,285.4 ppm BPA plus 285.4 ppm DBP,and a control diet,respectively,for 90 consecutive days.At the end of the study,the animals were sacrificed by exsanguination via the carotid artery under diethyl etherane aesthesia and weighed.Organs,including liver,kidneys,spleen,thymus,heart,brain,and testis underwent pathological examination.The androgen receptor（AR）,gonadotropin‐releasing hormone receptor（GNRHR）,and progesterone hormone receptor（PR） genes from the hypothalamus were detected by real‐time PCR.The biomedical parameters were analyzed.Results No significant difference was found in food intake,body weight,tissue weight,organ/brain weight ratio,and biomedical parameters among the four groups（P〉0.05）.However,BPA and DBP up‐regulated AR,PR and GNRHR expression levels in rats and showed a synergistic or an additive effect in the BPA＋DBP group.Conclusion The combined subchronic toxicity of BPA and DBP is synergistic or additive in male SD rats.

Effects of Extracellular ATP on Survival of Sensory Neurons in the Dorsal Root Ganglia of Rats

ATP was added to the cultured sensory neurons obtained from the dorsal root ganglia of the neonatal rats and PBS was added to serve as control. MTT assays were conducted to evaluate the survival and activity of the cultured neurons. And the silicone regenerative chamber was used after the sciatic nerve incision of the mature SD rat. 1 mmol/L ATP was injected into the left chamber and 0.09 % natrium chloride was injected into the right chamber as controls. The changes of nitric oxide synthase (NOS) activity in the corresponding dorsal root ganglia were measured histochemically and image analysis was also performed 4 days after the sciatic nerve injury. The results showed that extracellular ATP could enhance the survival of the neurons and the number of NOS positive neurons were significantly different between the ATP and control groups ( P <0.05). It was suggested that extracellular ATP had neurotrophic effect on neurons survival and could inhibit the NOS activity of the sensory neurons after the peripheral nerve incision, hence exerting the protective effect on the neurons, which was valuable for nerve regeneration after nerve injury.

Hyperbaric oxygen treatment induces dynamic ATPase activity changes in the rat brain following transient global cerebral ischemia-reperfusion

BACKGROUND： Energy depletion, induced by ischemia or hypoxia, is one of the first events in neuronal injury. OBJECTIVE： To investigate the dynamic changes of Na^＋-K^＋-ATPase and Ca^2＋-ATPase activity in the rat brain following transient global cerebral ischemia-reperfusion （IR）, as well as the effects of hyperbaric oxygen （HBO） treatment. DESIGN, TIME AND SETTING： A randomized and controlled animal study was performed in the Department of Biochemistry and Molecular Biology, Capital Medical University between February and December 2006. MATERIALS： Clean-grade, female, Sprague Dawley rats were provided by the Animal Research Department of Capital Medical University （License number： SYXK11-00-0047）. Na^＋-K^＋-ATPase and Ca^2＋-ATPase kits were provided by Nanjing Jiancheng Bioengineering Institute （Nanjing, China）. A hyperbaric oxygen chamber （DWC150-300） was supplied by Shanghai 701 Medical Oxygen Chamber Factory （Shanghai, China）. METHODS： Sixty-three rats were randomly divided into nine groups： sham operated group （sham-O） as control, groups of IR, and groups treated with hyperbaric oxygen （HBO） after IR. Animal from the IR and HBO groups were sacrificed after four different survival intervals of 6, 24, 48 and 96 hours, respectively. Each group consisted of seven rats. The rats of HBO groups were placed into the hyperbaric chamber. The HBO chamber was flushed with pure oxygen for 5 minutes, followed by a gradual rise in pressure over 5 minutes and stabilization at 0.2 MPa. Then, pure oxygen was supplied for 45 minutes in stabilized pressure, followed by gradually reduced pressure over 15 minutes. The rats of the 6-h HBO group were placed into the HBO chamber following reperfusion for 3 hours on the first day, which was repeated on three consecutive days, always at the same time. Rats in the sham-O group and IR group remained under normal atmospheric pressure. MAIN OUTCOME MEASURES： The Na^＋-K^＋-ATPase and Ca^2＋-ATPase activity in rat brain homogenate was detected by the ammonium molybdate assay method. RESULTS： All 63 rats were included in the final analysis. Alter 6 hours, Na^＋-K^＋-ATPase activity was significantly greater in HBO animals, compared with IR animals （P 〈 0.05） and sham-O controls （P〈 0.01）. In both, the HBO group and IR group, Na^＋-K^＋-ATPase activity returned to normal levels after 24 hours （P 〉 0.05）. At 48 and 96 hours, Na^＋-K^＋-ATPase activity was significantly greater in HBO and IR animals, compared with sham-O animals （P 〈 0.05）. Ca^2＋-ATPase activity was significantly greater in the HBO group after 6 hours, compared with the sham-O group （P 〈 0.01 ）, and returned to normal levels at 24 and 96 hours （P 〉 0.05）. In the IR group, Ca^2＋-ATPase activity was significantly higher after 6 hours than in the sharn-O group （P〈 0.01）, and returned to normal levels after 24 hours （P 〉 0.05）. CONCLUSION： The Na^＋-K^＋-ATPase and Ca^2＋-ATPase activity in IR groups increased during the acute and the delayed phase following transient global cerebral IR. HBO treatment not only increased Na^＋-K^＋-ATPase activity at the acute stage, but also induced a faster recovery of Ca^2＋-ATPase activity.

Effects of diethyldithiocarbamate on myelin basic protein expression in the rat lateral olfactory tract

BACKGROUND： Dithiocarbamates can cause demyelination of axons in the peripheral nervous system. Its derivate, diethyldithiocarbamate, is cytotoxic, and causes olfactory mucosal damage and atrophy of the olfactory bulb. However, it is still unclear whether the myelin sheath of the lateral olfactory tract is affected by diethyldithiocarbamate. OBJECTIVE： To investigate the effects of diethyldithiocarbamate on the myelin sheath of the rat lateral olfactory tract. This was done by examining changes in myelin basic protein expression after diethyldithiocarbamate treatment. DESIGN, TIME AND SETTING： A randomized, controlled, animal study was performed at the Laboratory of the Department of Human Anatomy and Neurobiology, Xiangya School of Medicine, Central South University, China from July to November 2007. MATERIALS： A total of 72 Sprague Dawley rats were randomly assigned into a diethyldithiocarbamate group （n = 32）, a solvent control group （n = 32）, and a blank control group （n = 8）. The diethyldithiocarbamate and solvent control groups were separately divided into 3-d, 7-d, 14-d and 28-d survival subgroups, with eight rats in each. Diethyldithiocarbamate （Sigma, USA） and goat anti-myelin basic protein polyclonal antibody （Santa Cruz, USA） were used in this study. METHODS： Rats in the diethyldithiocarbamate and solvent control groups were subcutaneously injected with diethyldithiocarbamate （600 mg/kg） and 0.01 mol/L phosphate buffered saline （600 mg/kg） at the posterior neck, respectively. Rats in the blank control group received no treatment. MAIN OUTCOME MEASURES： Immunohistochemical staining and Western blot assay were used to measure myelin basic protein expression in the rat lateral olfactory tract. RESULTS： Following immunohistochemical staining, myelin basic protein was uniformly distributed in the rat lateral olfactory tract in the blank control and solvent control groups. Western blot assay showed 21.5, 18, 17 and 14 ku positive bands. No significant difference was found in myelin basic protein distribution and blot pattern, in the rat lateral olfactory tract, in the diethyldithiocarbamate group, following immunohistochemical staining and Western blot assay. Myelin basic protein expression gradually decreased at day 3, reached the lowest level at day 7, and gradually increased again at days 14 and 28. CONCLUSION： Demyelination is induced by diethyldithiocarbamate in the rat lateral olfactory tract in an early stage, followed by remyelination at later stages.

Isolation of genes related to blood glucose-control in rat skeletal muscle

Obejctive To isolate genes related to blood glucose control using Sprague Dawley (SD) rat skeletal muscle Methods Differential gene expression between glucose stimulated and non glucose stimulated SD rat skeletal muscle was obtained by the differential display (DD) method, Slot blotting hybridization and Northern blot hybridization Results Several new genes that are differentially expressed in glucose stimulated and non glucose stimulated SD rat skeletal muscle were isolated 74 were verified by slot analysis from 181 gene tags isolated Of them, 33 were cloned and sequenced, and homologous analysis and application for GenBank Access Number were carried out 21 expressed sequence tag (EST) representing novel genes was confirmed by Northern blot analysis A total of 9 novel genes showed significant differential expression patterns Conclusions Using improvements and modifications of the differential display technique, a labor and cost saving route was used to identify new genes related to blood glucose control We investigated differentially expressed genes at the whole body level instead of the culture cell level, to ensure experimental results closer to the normal physiological state This technique may be valid in wide spread application to other related research