Capsaicin restores gubernacular contractility in TS rats

BACKGROUND/PURPOSE: Calcitonin gene-related peptide (CGRP) may stimulate gubernacular migration during testicular descent by release from the genitofemoral nerve (GFN). The origin of CGRP within the nerve, however, is controversial. This study examines whether sensory nerve destruction alters gubernacular contractility in vitro in Sprague-Dawley (SD) and congenitally cryptochid (TS) rats. METHODS: Part 1: Twenty-four SD and 16 TS rats (day 0) had either both GFNs transected or sham operation. Gubernacula were removed on day 2 and cultured with or without CGRP (714 nmol/L). Contractility was recorded by video. Part 2: Twenty-two SD and 17 TS rats (day 0) were injected with either capsaicin or vehicle. Gubernacula were removed (day 2) and cultured as above. RESULTS: Part 1: In sham-operated SD rats gubernacular contracility increased from 8% to 83% with added CGRP. After GFN transection contractility was not affected by CGRP (21% without and 86% with CGRP; not significant). TS rat gubernacula had no endogenous contractions, but after GFN transection, the contractile response to CGRP increased from 6% to 44% (P = .04). Part 2: In vehicle-treated SD rats, rhythmic contractions increased from 10% to 86% with CGRP, which was unchanged by capsaicin treatment (82%; not significant). In vehicle-treated TS rats, gubernacular contractions were 6% after CGRP. After capsaicin pretreatment, contractions increased to 59% with CGRP (P = .002). CONCLUSIONS: Results of this study show that chemical destruction of sensory nerves restores gubernacular contractility in mutant cryptorchid TS rats. Release of CGRP appears to occur through sensory nerves.     

Undescended testis is accompanied by calcitonin gene related peptide accumulation within the sensory nucleus of the genitofemoral nerve in trans-scrotal rats

PURPOSE: Recent data suggest that calcitonin gene related peptide (CGRP) released from the sensory branch of the genitofemoral nerve may regulate testicular descent. We studied the number of CGRP immunoreactive cells in the sensory nucleus of the genitofemoral nerve (L1 to L2 dorsal root ganglia) in cryptorchid trans-scrotal rats. Four-week-old trans-scrotal rats with unilateral undescended testis underwent bilateral genitofemoral nerve dissection and retrograde nerve labeling with the fluorescent dye 4,6-diamidino-2-phenylindole (DAPI). Animals were sacrificed 48 hours later and the L1 to L2 dorsal root ganglia were removed. Serial sections were obtained and double fluorescent labeled with antibody to CGRP. Retrograde labeled and CGRP immunoreactive cells were counted using an epi-fluorescent microscope.In the 6 male trans-scrotal rats evaluated we noted a mean plus or minus standard deviation of 1,272 +/- 98 retrograde labeled dorsal root ganglion cells ipsilateral to a fully descended testicle, including 98 +/- 34 that were also CGRP immunoreactive. On the side of the undescended testis there was a mean of 1,600 +/- 304 DAPI positive cells and 160 +/- 51 CGRP immunoreactive, DAPI labeled cells. The difference was significant (p <0.02).This study shows that in trans-scrotal rats the sensory nucleus of the genitofemoral nerve contains more CGRP immunoreactive cells ipsilateral to an undescended testis than on the contralateral side, highlighting the significance of CGRP supply through the sensory branch of the genitofemoral nerve for testicular descent.     

Exogenous calcitonin gene-related peptide can change the direction of gubernacular migration in the mutant trans-scrotal rat.

PURPOSE: The mutant trans-scrotal (TS) rat shows unilateral or bilateral suprainguinal testes in more than 70% of males. Calcitonin gene-related peptide (CGRP) has been proposed as the neurotransmitter released from the genitofemoral nerve (GFN), which controls gubernacular migration to the bottom of the scrotum during inguinoscrotal descent. Results of previous studies in this rat suggest a down-regulation of CGRP receptors in gubernaculum occurring caused by excess release of the neuropeptide from the GFN. The aim of this study was to test the hypothesis that division of the GFN in neonatal TS rats, potentially allowing the gubernaculum to become sensitive to CGRP, followed by exogenous CGRP injections, would change the direction of gubernacular migration and the final position of the testis. METHODS: Four study groups were used: group 1 (n = 43), sham operation, in which the peritoneal cavity was opened and the left genitofemoral nerve was exposed but not divided, and oil injections into left hemiscrotum; group 2 (n = 70), division of left GFN and CGRP injections into left hemiscrotum; group 3 (n = 36), sham operation and CGRP injections into left hemiscrotum; group 4 (n = 30), division of left GFN and oil injections into left hemiscrotum. RESULTS: In group 2 (GFN division and CGRP injection), 18 testes were located in a position not previously described in this model. In 16 (23%) rats, the testis was located at the entrance of the internal inguinal ring with the gubernaculum directed down toward the scrotum. This contrasts with the normal position of the testis in the superficial inguinal pouch, where the testis is located superficial to the external oblique muscle, with the gubernaculum directed craniolaterally. In two (3%) rats, there was incomplete descent of the testis. In group 3 (sham operation and CGRP injection), two (6%) testes were located at the internal ring. The remaining testes in the above groups and all of the testes in groups 1 and 4 were found in either the superficial inguinal pouch, completely descended, or adherent to scar tissue. CONCLUSIONS: These findings suggest that division of the GFN in neonatal TS rats followed by CGRP injections into the scrotum can change the direction of gubernacular migration in the TS rat. The effectiveness of this experimental model is limited by the inability to accurately localize injected CGRP into the scrotum.     

THE GENITOFEMORAL NERVE MAY LINK TESTICULAR INGUINOSCROTAL DESCENT WITH CONGENITAL INGUINAL HERNIA

The genitofemoral nerve (GFN) hypothesis for inguinoscrotal testicular descent proposes that calcitonin gene-related peptide (CGRP), released from the genitofemoral nerve, controls the migration of the gubernaculum from the inguinal region to the scrotum between 26 and 40 weeks of gestation. The processus vaginalis provides a channel through which the testis descends from the abdomen to the scrotum. Following descent of the testis the processus vaginalis undergoes luminal obliteration and disappearance between the internal inguinal ring and the upper pole of the testis. The mechanism underlying closure of the processus is unknown and failure for it to occur normally results in congenital inguinal hernia, scrotal hydrocele and possibly even an ‘ascending’ testis. Recent work in our laboratory suggests that CGRP, released from the genitofemoral nerve, may cause fusion and disappearance of the processus vaginalis. We propose that abnormalities in the GFN link a spectrum of disorders encompassing congenital undescended testis, inguinal hernia, scrotal hydrocele and ascending testis.     

荧光逆行示踪法定位神经端侧缝合后再生来源的实验研究

目的 应用荧光逆行示踪法研究神经端.侧缝合修复臂丛神经损伤的有效性及再生神经的脊髓定位.方法 雌性SD大鼠24只,随机分为4组,造成臂丛神经上干损伤模型,分别以膈神经、同侧颈,神经根为供体神经,按照端.侧和端.端两种缝合方式修复肌皮神经.术后3个月,对大鼠肌皮神经和供体神经分别采用真蓝和双脒基黄进行逆行示踪.3、7、14 d后进行灌注固定,取颈段脊髓连续切片,荧光显微镜观察.结果 各观察点背根节及脊髓前角均出现荧光标记细胞,并逐渐增多.以同侧颈,为供体神经组,标记细胞仅出现在该节段,而以膈神经为供体神经组,标记细胞出现在颈_(3-5)节段.端一侧缝合组在相应脊髓前角或背根神经节出现,同时具有两种荧光剂的双标细胞或在同一脊髓节段同时出现分别以两种荧光剂标记的单标细胞.结论 采用不同供体神经进行端.侧缝合联合神经移植修复臂丛神经可使神经再生,荧光逆行示踪可以准确定位端.侧缝合后再生神经的来源.     

大鼠皮肤降钙素基因相关肽正常分布及烫伤后改变规律

目的 探讨皮肤创面降钙素基因相关肽（ＣＧＲＰ）的来源。方法 运用免疫组化技术检测烫伤后早期大鼠皮肤烫伤创面,创周及远处未损伤皮肤内含ＣＧＲＰ的神经分布密度改变。结果 烫伤后１５ｍｉｎ在以上所有部位的含ＣＧＲＰ神经纤维分布密度明显下降,烫伤后６至１２ｈ达到最低值,而后逐渐恢复,相比之下,在创周恢复过程出现较早;此外,在创面和创周的真皮层ＣＧＲＰ免疫反应阳性的巨噬细胞样大细胞从局部血管中游出,烫伤后１     

Possible mechanism of referred pain in the perineum and pelvis associated with the prostate in rats

PURPOSE: Since persistent pain in the perineum and pelvic floor associated with chronic prostatitis /chronic pelvic pain syndrome has been hypothesized to be referred pain, it might also be explained by neural mechanisms. MATERIALS AND METHODS: Dual retrograde fluorescent labeling and immunohistochemistry were identified as methods with which to investigate the neurogenic aspect of this status. The dual distribution of dorsal root ganglia (DRG) cells was determined after double retrograde fluorescent staining of the prostate and pelvic floor, and the prostate and perineum somatic nerves. Calcitonin gene-related peptide (CGRP) and substance P (SP) in dual labeled cells were determined by immunohistochemistry, giving possible insight into the cause of pelvic pain. RESULTS: Fluorescent double labeled cells were found in the lumbar and sacral DRG, while double labeled cells were distributed predominantly in L6 to S1 and L1 to L2 segment DRG in groups 1 and 2, respectively. On immunohistochemistry some of them were confirmed to contain CGRP and SP. Thus, there are crossover pathways between the prostate and pelvic floor. CONCLUSIONS: The findings that we present confirm that the peripheral process of DRG cells dichotomizes to the prostate, sphincter and somatic parties simultaneously. Some of these cells contain CGRP and SP, which indicate that referred pain in the perineum and pelvic floor may be caused by an axon reflex in the peripheral process of DRG neurons.     

Pathology of lumbar nerve root compression. Part 2: morphological and immunohistochemical changes of dorsal root ganglion.

STUDY DESIGN: This study is to investigate the changes of dorsal root ganglion (DRG) induced by mechanical compression using in vivo model. OBJECTIVES: The effect of axonal flow disturbance induced by nerve root compression was determined in DRG. SUMMARY OF BACKGROUND DATA: The dorsal root ganglion should not be overlooked when considering the mechanism of low back pain and sciatica, so it is important to understand the morphologic and functional changes that occur in primary sensory neurons of the dorsal root ganglion as a result of nerve root compression. However, few studies have looked at changes of neurons within the dorsal root ganglion caused by disturbance of axonal flow and the axon reaction as a result of mechanical compression of the dorsal root through which the central branches of the primary sensory nerves pass. METHODS: In mongrel dogs, the seventh lumbar nerve root was compressed for 24 h, one week, or three weeks using a clip with a pressure of 7.5 gf. Morphologic changes of the primary sensory neurons in the dorsal root ganglion secondary to the axon reaction were examined by light and electron microscopy. Changes of immunostaining for substance P (SP), calcitonin gene-related peptide (CGRP), and somatostatin (SOM) in the primary sensory neurons affected by central chromatolysis after nerve root compression were also examined. RESULTS: Light microscopy showed central chromatolysis of neurons in the dorsal root ganglion from one week after the start of compression. Electron microscopy of the affected neurons revealed movement of the nucleus to the cell periphery and the loss of rough endo-plasmic reticulum and mitochondria from the central region. Immunohistochemical studies showed a marked decrease of SP, CGRP, and SOM staining in small ganglion cells with central chromatolysis when compared with cells from control ganglia. CONCLUSION: It is important to be aware that in patients with nerve root compression due to lumbar disc herniation or lumbar canal stenosis, dysfunction is not confined to degeneration at the site of compression, but also extends to the primary sensory neurons within the dorsal root ganglion as a result of the axon reaction. Patients with sensory disturbance should therefore be fully informed of the fact that these symptoms will not resolve immediately after surgery.     

大鼠烫伤后肾上腺内含降素基因相关肽神经的改变

目的 探讨皮肤烫伤早期大鼠肾上腺内降钙素基因相关肽（CGRP）变化规律,为阐明烫伤后肾上腺内分泌功能改变的机理提供资料。方法 采用15％深Ⅱ度大鼠烫伤模型,以免疫组化方法分别检测烫伤后不同时相点肾上肾皮质、髓质CGRP免疫反应阳性神经和细胞分布密度的改变。结果 ①在肾上腺被膜,皮质,髓质均有含CGRP神经分布,其中髓质更为密集。在髓质可见含CGRP神经与CGRP免疫反应阳性细胞密切接触;②烫伤早期大鼠肾上腺皮质和髓质含CGRP神经的分布密度均下降,但髓质内CGRP免疫反应阳性细胞的分布密度上升。结论 降钙素基因相关肽可能是烫伤大鼠肾上腺功能改变的影响因素之一。     

Calcitonin gene-related peptide partially mediates nociception in acute experimental pancreatitis

BACKGROUND: The mechanism by which pancreatitis causes pain is unknown. The neuropeptide calcitonin gene-related peptide (CGRP) is released after sensory nerve activation and promotes nociceptive signaling in models of visceral pain. We hypothesized that acute pancreatitis leads to the activation of pancreatic sensory neurons that release CGRP in the dorsal horn of the spinal cord. This signal is ultimately transmitted to the brain, and pain is sensed. METHODS: To induce pancreatitis, rats were injected with l-arginine (500 mg/kg) intraperitoneally or saline (control). Pancreatitis was confirmed by measuring serum amylase and evaluating pancreatic histology. Activation of nociceptive pathways was evaluated by counting Fos-like immunoreactive nuclei (FLI) in the dorsal horn of the spinal cord at T3-L1. Some animals received the CGRP antagonist CGRP(8-37) (50 microg intrathecally) 2 hours before perfusion. Animals were compared using a 2-tailed t test. RESULTS: l-Arginine treatment induced acute necrotizing pancreatitis in the rat at 24 hours. l-Arginine (24 hours) increased FLI in the dorsal horn of the spinal cord, with a peak effect at L1. Intrathecal administration of CGRP(8-37) significantly decreased the number of FLI nuclei in the dorsal horn of the spinal cord in T11-L1. CONCLUSIONS: Nociception in the l-arginine model of acute pancreatitis is partially mediated by the release of CGRP in the dorsal horn of the spinal cord. Antagonism of CGRP or its receptors may be useful in treating pain from acute pancreatitis.     

磷脂酶A2硬膜外腔注射对大鼠腰髓及背根节中CGRP含量的影响

目的探讨腰椎间盘突出症相关腰腿痛发病的可能机制.方法在大鼠的硬膜外腔注射磷脂酶A2,免疫组化的方法测定大鼠背根节和脊髓后角中CGRP的变化.结果硬膜外腔注射磷脂酶A2后,L4-6背根节中CGRP阳性神经元的数目和面积明显增加,脊髓背角浅层中CGRP阳性神经纤维终末的面积也明显增加.证实与对照组相比较有显著性差别(P＜0.05).结论磷脂酶A2是一种疼痛伤害性刺激,它可能在腰椎间盘突出相关腰腿痛的发病中起到重要的作用.     

神经病理性痛大鼠背根神经节和脊髓背角Nogo-A蛋白表达的变化

目的 评价神经病理性痛大鼠背根神经节和脊髓背角Nogo-A蛋白表达的变化.方法 健康成年雄性SD大鼠72只,体重250～300 g,采用随机数字表法,将大鼠随机分为3组(n=24):对照组(C组)、假手术组(S组)和神经病理性痛组(NP组).C组不做任何处理,NP组采用结扎并剪断胫神经和腓总神经的方法制备大鼠神经病理性痛模型,S组仅切皮暴露坐骨神经但不结扎和剪断神经.于结扎后1、7、14、21 d时采用yon Frey丝测定大鼠机械痛阈.于各时点随机取6只大鼠处死后取损伤侧L5背根神经节和L4,5脊髓,采用免疫荧光标记法测定Nogo-A蛋白的表达(n=3),采用Westernblot法测定Nogo-A蛋白的表达(n=3).结果 与C组和S组比较,NP组大鼠结扎后7、14、21 d时机械痛阈降低,结扎后7、14 d时背根神经节Nogo-A蛋白表达下调,结扎后14、21 d时脊髓背角Nogo-A蛋白表达上调(P＜0.05).结论 背根神经节及脊髓背角Nogo-A蛋白可能在外周神经损伤诱发大鼠神经病理性痛过程中发挥重要作用.

Abstract：

Objective To investigate the changes in the expression of Nogo-A protein in the dorsal root ganglion (DRG) and spinal dorsal horn in a rat model of neuropathic pain (NP) .Methods Seventy-two male SD rats weighing 250-300 g were randomly divided into 3 groups ( n = 24 each) : control group (group C) , sham operation group (group S) and NP group. NP was induced by ligation and severance of tibial and common fibular nerves according to the technique described by Isabelle et al. The mechanical withdrawal threshold (MWT) to von Frey filament stimulation was measured at 1, 7, 14 and 21 days after ligation. Six rats in each group were randomly selected at each time point and sacrificed (3 for determination of Nogo-A protein expression by immunofluorescence, 3 for determination of Nogo-A protein expression by Western blot) . The L5 DRG and L4,5 segment of spinal cord on the injured side were removed for determination of Nogo-A protein expression by immunofluorescence and Western blot. Results Compared with the groups C and S, MWT was significantly decreased at 7, 14 and 21 days after ligation, the expression of Nogo-A protein in the DRG was down-regulated at 7 and 14 days after ligation and the expression of Nogo-A protein in the spinal dorsal horn was up-regulated at 14 and 21 days after ligation ( P ＜0.05) .Conclusion The Nogo-A protein in the DRG and spinal dorsal horn may play an important role in peripheral nerve injury-induced NP in rats.     

Pubertal genitofemoral nerve division induces testicular ascent in adult rats

BACKGROUND/PURPOSE: Ascending testes, which normally are located at the bottom of the scrotum in early infancy and later ascend back out of the scrotum, have been reported by several investigators. However, little is known about the effect of the division of the genitofemoral nerve (GFN) on testicular ascent as boys grow. The purpose of this study is to investigate whether the division of the proximal genitofemoral nerve in prepubertal rats induces testicular ascent in adulthood. METHODS: Thirty-day-old Wistar King A Rats (n = 27) underwent a unilateral proximal GFN transection on either the right or left side. At 150 days of age, the rats were killed, and their testicular position was examined. The length of the processus vaginalis was measured, and the testes were removed and weighed. Sham-operated rats were used as controls (n = 10). Student's t and the chi2 test were used for the statistical analysis. RESULTS: At 150 days of age, 21 of the 27 operated rats (77.8%) showed unilateral testicular ascent on the operated side. All testes were located at the bottom of the scrotum in sham-operated control rats (20 testes). Both the length of the processus vaginalis and the testicular weight were decreased significantly more on the operated side than in the sham-operated rats. CONCLUSIONS: These findings suggest that the proximal division of the genitofemoral nerve in prepubertal rats may induce a relative ascent of the testis by preventing the growth of the processus vaginalis in adulthood. In patients with such ascending testes, an abnormal development or accidental trauma of the genitofemoral nerve may be involved in testicular ascent.     

大鼠皮肤降钙素基因相关肽正常分布及烫伤后改变规律

目的探讨皮肤创面降钙素基因相关肽(CGRP)的来源。方法运用免疫组化技术检测烫伤后早期大鼠皮肤烫伤创面,创周及远处未损伤皮肤内含 CGRP 的神经分布密度改变。结果烫伤后15 min 在以上所有部位的含 CGRP 神经纤维分布密度明显下降。烫伤后6至12 h 达到最低值,而后逐渐恢复,相比之下,在创周恢复过程出现较早;此外,在创面和创周的真皮层 CGRP免疫反应阳性的巨噬细胞样大细胞从局部血管中游出,烫伤后12 h 该细胞与含 CGRP 神经关系密切,烫伤后24 h 该细胞染色增强,破碎并释放大量 CGRP 免疫反应阳性的颗粒状物质,而后这些细胞在局部消失。结论在皮肤创面针对烧伤损害 CGRP 可能不仅从皮肤神经末端释放,也能由局部炎性细胞合成释放。     

Androgen and estrogen receptor expression in the spinal segments of the genitofemoral nerve during testicular descent

Aim

During testicular descent (TD), the genitofemoral nerve (GFN) is masculinized by androgen. This study aimed to test whether androgen receptor (AR), estrogen receptorα (ERA), or estrogen receptor β (ERB) are expressed during TD in the GFN spinal segments and dorsal root ganglia (DRG) in normal and flutamide-treated rats.

Methods

Time-mated Sprague-Dawley dams were injected with flutamide (75 mg/kg, subcutaneously (S/C) in sunflower oil) on embryonic (E) days 16 to 19. Embryonic and postnatal (P) male L1-2 spinal cord segments were collected (E16, E17, E19, P0, P2, and P4) in control and flutamide-treated groups (n = 5-10). Samples were fixed in 4% paraformaldehyde. Five-micrometer-thick sections were prepared immunohistochemically for AR, ERA, and ERB.

Results

During TD, ERB was expressed in L1-2 DRG. Surprisingly, AR was not expressed in prenatal DRG, only after P2. There was no ERA expression. Flutamide had no effect on AR, ERB, or ERA expression in the L1-2 DRG during TD.

Conclusion

During the E window of androgen sensitivity, the GFN is not directly masculinized, with little AR expression and no change with flutamide over this period. Estrogen receptor β is expressed in the DRG during TD. However, its relevance is yet to be determined.     

鞘内注射舒芬太尼对神经病理性痛大鼠脊髓背角NMDA受体及CGRP表达的影响

目的 评价鞘内注射舒芬太尼对神经病理性痛大鼠脊髓背角N-甲基-D-天冬氨酸(NMDA)受体及降钙素相关基因肽(CGRP)表达的影响.方法 雄性SD大鼠36只,体重220～280 g,随机分为4组(n=9):正常对照组(C组)、假手术组(S组)、坐骨神经分支选择性损伤组(SNI组)和舒芬太尼+坐骨神经分支选择性损伤组(S+SNI组).SNI组和S+SNI组制备SNI模型,S+SNI组在SNI术后14 d内每天鞘内注射舒芬太尼1 μg(用生理盐水稀释至10 μl),其余各组给予等容量生理盐水.于SNI给药前2 d(基础状态)及给药1、2、7、14 d测定机械痛阈和热缩足潜伏期,分别于给药2、7、14 d测定痛阈后立即处死3只大鼠,采用免疫组化法测定L5节段脊髓背角NMDA受体和CGRP表达水平.结果 与C组和S组比较,SNI组机械痛阚降低,NMDA受体和CGRP表达上调(P<0.01),热缩足潜伏期差异无统计学意义(P>0.05).与SNI组比较,S+SNI组机械痛阈升高,热缩足潜伏期延长,NMDA受体和CGRP表达下调(P<0.01).结论 鞘内注射舒芬太尼可抑制脊髓背角NMDA受体和CGRP表达上调,从而减轻大鼠神经病理性痛.     

周围神经端端或端侧吻合后降钙素基因相关肽和P物质免疫组织化学的对比研究

目的　探讨周围神经端端或端侧吻合后神经递质降钙素基因相关肽 (CGRP)和 P物质 (SP)水平的变化。　方法　选取雌性 Wistar大鼠 2 0只 ,随机分为 5组 ,每组 4只 ;实验组为 4组 ,正常对照组为 1组。实验组 :切断双侧腓总神经。将左侧腓总神经远侧断端吻合至胫神经侧方 ,右侧行腓总神经端端吻合 ,术后 1、2、4和 2 7周于各时间点分别在神经吻合口、腰段脊髓和背根神经节处取材 ,每次 4只进行 CGRP和 SP的免疫组织化学染色。对照组 :不行神经切断及吻合术 ,1周后检测同上。　结果　实验组术后 1周在腰段脊髓后角和背根神经节中 CGRP和 SP的表达均显著减少 ,术后 4、2 7周 ,CGRP和 SP在腰段脊髓后角的表达逐渐恢复至接近正常水平 ,但在背根神经节中的表达却无显著变化。端端、端侧吻合的两侧 CGRP和 SP变化的趋势一致 ,但在腰段脊髓后角的 4个时间点的样本中 ,端端吻合的表达显著高于端侧吻合。对坐骨神经进行乙酰胆碱酯酶 (Ach E)、SP、CGRP和 PGP9.5染色显示神经纤维均可通过端端与端侧神经吻合口。　结论　再生的神经纤维可以通过端侧吻合口 ,端侧吻合中感觉神经的恢复与端端吻合相似 ,但恢复的程度稍差于端端吻合。     

Characterization of the induced neuropeptide Y-like immunoreactivity in primary sensory neurons following complete median nerve transection

In this study, we examined characteristics of the neuropeptide Y-like immunoreactive (NPY-LI) dorsal root ganglion (DRG) neurons after complete median nerve transection (CMNT). With fluorogold (FG) injection into normal median nerves, numerous FG-labeled DRG neurons were localized predominantly in the C6 and C7 DRGs, where the focal regions were examined after CMNT. With NPY immunohistochemistry, a few NPY-LI neurons were detected in the ipsilateral but not contralateral DRGs after FG injection into the nerve. As early as 3 days after CMNT a few NPY-LI neurons could be detected, reaching a maximum in the DRGs at 4 weeks, subsiding thereafter over 20 weeks. The NPY-LI DRG neurons were primarily medium-sized and large neurons. With FG injection into the transected median nerve, we found that approximately 99% of NPY-LI neurons were labeled for FG, suggesting that they were derived from the injured but not intact DRG neurons. Using double fluorescent dyes tracing, we detected that some of the injured DRG neurons were NPY-LI neurons that projected to the cuneate nucleus (CN). Following dorsal rhizotomy, our data indicated that after CMNT the induced NPY-LI fibers in the ipsilateral CN originated exclusively from the injured DRG neurons. Taken together, these findings suggest that injury-induced NPY-LI fibers in the CN may originate from the injured DRG neurons via the median primary afferent fibers, affect the excitability of cuneothalamic projection neurons (CTNs), and involve neuropathic sensation following CMNT.     

Pancreatic tissue grafts are reinnervated by neuro-peptidergic and cholinergic nerves within five days of transplantation

The reinnervation process is crucial for the survival and functioning of cell, tissue or organ transplants. This study was designed to examine the exact time of reinnervation of intraocular pancreatic tissue transplants in rats. The rate of survival of neuropeptide-containing cells in pancreatic tissue grafts was also investigated. Calcitonin gene-related peptide (CGRP), galanin (GAL), neuropeptide Y (NPY) were observed in the surviving nerve cell bodies of the grafts. The iridal nerves reinnervating the pancreatic grafts expressed CGRP, GAL, NPY and choline-acetyl-transferase (ChAT) on day 5, and tyrosine hydroxylase (TH) and nitric oxide synthase (bNOS) on day 6 of the transplantation period. The expression of CGRP in the reinnervating nerves was more consistent when compared to GAL, NPY, ChAT, TH and bNOS. Although all of the three neuropeptides (CGRP, GAL, NPY) were present in the surviving nerve cell bodies of the pancreatic tissue graft up to the end (day 9) of the transplantation period, the number of CGRP-immunopositive cells was consistently higher throughout the transplantation period. Hence, the number of CGRP-positive cells in the pancreatic tissue graft was significantly (P < 0.05) higher than that of GAL and NPY. In conclusion, pancreatic fragments were reinnervated by neuropeptidergic (CGRP, NPY) and cholinergic (ChAT) nerves within the first 5 days of transplantation. In addition to the reinnervation of pancreatic tissue grafts, the intrinsic neurones of the grafts also survived after transplantation. The rate of survival of CGRP-containing cells in the pancreatic tissue grafts was more consistent compared to that of NPY and GAL.     

Antihyperalgesic Effect of a Recombinant Herpes Virus Encoding Antisense for Calcitonin Gene-related Peptide

Background: Calcitonin gene-related peptide (CGRP) is contained in and released by small-diameter, nociceptive primary afferent sensory neurons. Upon spinal release, one of the effects of CGRP seems to be to sensitize dorsal horn neurons to subsequent input from nociceptive afferents and, consequently, to induce a behavioral hyperalgesia. Therefore, attenuating evoked release of CGRP from central terminals of nociceptors should have an antihyperalgesic effect.

Methods: The authors applied a recombinant herpes vector, encoding an antisense sequence to the whole CGRP gene, to the dorsal surface of the hind paw of mice to knock down expression of the peptide selectively in primary afferents innervating this tissue.

Results: Herpes virus-based vector encoding an antisense sequence for the whole CGRP clearly reduced CGRP immunoreactivity in the infected spinal dorsal horn levels as well as in cultured dorsal root ganglia neurons. Selective knockdown of CGRP in primary afferents significantly attenuated the thermal, C-fiber hyperalgesia normally observed after topical application of capsaicin. The effect of viral vector-mediated knockdown of CGRP was comparable to the effect of intrathecal application of the CGRP antagonist CGRP8-37, but lasted for 14 weeks after one single application.