Experimental model for controlled endoscopic subepithelial vocal fold injury in rats

Purpose: To present a detailed, reproducible, cost-efficient surgical model for controlled subepithelial endoscopic vocal fold injury in the rat model. Methods: Six male Sprague Dawley rats were enrolled in the experiment. The left vocal folds were used to carry out the injury model, and the right vocal fold served as control. After deep sedation, the rats were placed on a custom operating platform. The vocal fold injury by subepithelial stripping was carried out using custom-made microsurgical instruments under endoscopic guidance. Data were analyzed for procedural time and post-procedural pain. Microcomputed tomography (micro-CT) scan and histologic images were obtained to assess the length, area, and depth of injury to the vocal fold. Results: The mean procedural time was 112 s. The mean control vocal fold length was 0.96 ± 0.04 mm. The mean vocal fold injury length was 0.53 ± 0.04 mm. The mean vocal fold surface was 0.18 ± 0.01 mm2 with a mean lesion area of 0.05 ± 0.00 mm2. Mean vocal fold injury depth was 375.4 ± 42.8 µm. The lesion length to vocal fold length ratio was 0.55 ± 0.03, as well as lesion area to vocal fold surface area was 0.29 ± 0.02. Conclusions: Our described experimental vocal fold injury model in rats is found to be fast, safe, cost-efficient, and reproducible with a rapid learning curve.

Galectin-3 in septic acute kidney injury: a translational study.

BACKGROUND: Galectin-3 (Gal-3) is a pleiotropic glycan-binding protein shown to be involved in sepsis and acute kidney injury (AKI). However, its role has never been elucidated in sepsis-associated AKI (S-AKI). We aimed to explore Gal-3's role and its potential utility as a therapeutic target in S-AKI. METHODS: In 57 patients admitted to the intensive care unit (ICU) with sepsis, serum Gal-3 was examined as a predictor of ICU mortality and development of AKI. In a rat model of S-AKI induced by cecal ligation and puncture (CLP), 7-day mortality and serum Gal-3, Interleukin-6 (IL-6), and creatinine were examined at 2, 8, and 24 hours (h) post-CLP. Two experimental groups received the Gal-3 inhibitor modified citrus pectin (P-MCP) at 400 mg/kg/day and 1200 mg/kg/day, while the control group received water only (n = 18 in each group). RESULTS: Among 57 patients, 27 developed AKI and 8 died in the ICU. Serum Gal-3 was an independent predictor of AKI (OR = 1.2 [95% CI 1.1-1.4], p = 0.01) and ICU mortality (OR = 1.4 [95% CI 1.1-2.2], p = 0.04) before and after controlling for age, AKI, and acute physiology and chronic health evaluation (APACHE II) score. In the CLP rat experiment, serum Gal-3 peaked earlier than IL-6. Serum Gal-3 was significantly lower in both P-MCP groups compared to control at 2 h post-CLP (400 mg: p = 0.003; 1200 mg: p = 0.002), and IL-6 was significantly lower in both P-MCP groups at all time points with a maximum difference at 24 h post-CLP (400 mg: p = 0.015; 1200 mg: p = 0.02). In the Gal-3 inhibitor groups, 7-day mortality was significantly reduced from 61% in the control group to 28% (400 mg P-MCP: p = 0.03) and 22% (1200 mg P-MCP: p = 0.001). Rates of AKI per RIFLE criteria were significantly reduced from 89% in the control group to 44% in both P-MCP groups (400 mg: p = 0.007; 1200 mg: p = 0.007). CONCLUSIONS: This translational study demonstrates the importance of Gal-3 in the pathogenesis of S-AKI, and its potential utility as a therapeutic target.

Vacuum therapy ameliorates erectile dysfunction in bilateral cavernous nerve crush rats by inhibiting apoptosis and activating autophagy.

Postprostatectomy erectile dysfunction (pPED) remains a current problem despite improvements in surgical techniques. Vacuum therapy is clinically confirmed as a type of pPED rehabilitation. However, its underlying mechanisms are incompletely understood. Recently, autophagy and apoptosis were extensively studied in erectile dysfunction resulting from diabetes, senescence, and androgen deprivation but not in the context of pPED and vacuum therapy. Therefore, this study was designed to investigate the roles of autophagy and apoptosis in pPED and vacuum therapy. Twenty-four adult male Sprague-Dawley rats were randomly divided into three groups: the control group, bilateral cavernous nerve crush (BCNC) group, and BCNC + vacuum group. After 4 weeks of treatment, intracavernosal pressure was used to evaluate erectile function. Real-time quantitative polymerase chain reaction, western blot, and immunohistochemistry were used to measure the molecular expression. TdT-mediated dUTP nick-end labeling staining was used to assess apoptosis. Transmission electron microscopy was used to observe autophagosomes. After treatment, compared with those of the BCNC group, erectile function and cavernosal hypoxia had statistically significantly improved (P < 0.05). Apoptosis and the relative protein expression of B-cell lymphoma-2-associated X and cleaved Caspase3 were decreased (P < 0.05). Autophagy-related molecules such as phosphorylated unc-51-like autophagy-activating kinase 1 (Ser757) and p62 were decreased. Beclin1, microtubule-associated protein 1 light chain 3 A/B, and autophagosomes were increased (P < 0.05). Besides, the phosphatidylinositol 3-kinase/AKT/mammalian target of rapamycin signaling pathway, as a negative regulator of autophagy to some degree, was inhibited. This study revealed that vacuum therapy ameliorated pPED in BCNC rats by inhibiting apoptosis and activating autophagy.

Establishing a successful rat model of duodenal- jejunal bypass: A detailed guide.

Gastric bypass surgery, an operation that restricts the stomach and bypasses the duodenum and part of the jejunum, results in major improvement or remission of type 2 diabetes. Duodenual-jejunal bypass was developed by one of the authors (FR) as an experimental, stomach-sparing variant of gastric bypass surgery to investigate weight-independent mechanisms of surgical control of diabetes. Duodenual-jejunal bypass has been shown to improve various aspects of glucose homeostasis in rodents and in humans, thus providing an experimental model for investigating mechanisms of action of surgery and elusive aspects of gastrointestinal physiology. Performing duodenual-jejunal bypass in rodents, however, is associated with a steep learning curve. Here we report our experience with duodenual-jejunal bypass and provide practical tips for successful surgery in rats. Duodenual-jejunal bypass was performed on 50 lean rats as part of a study aimed at investigating the effect of the procedure on the physiologic mechanisms of glucose homeostasis. During the study, we have progressively refined details of anatomic exposure, technical aspects of duodeno-jejunostomy and peri-operative care. We analysed the role of such refinements in improving operative time and post-operative mortality. We found that refinement of exposure methods of the gastro-duodenal junction aimed at minimizing tension on small visceral vasculature, technical aspects of duodeno-jejunal anastomosis and peri-operative management played a major role in improving the survival rate and operative time. Overall, an experimental model of duodenual-jejunal bypass was successfully reproduced. Based on this experience, we describe here what we believe are the most important technical tips to reduce the learning curve for the procedure.

Surgical Anatomy of Rats for the Training of Microvascular Anastomosis.

BACKGROUND: Microvascular anastomosis is an essential procedure in neurosurgery, but the opportunity to perform the surgery has gradually decreased for neurosurgeons. Therefore, training is necessary for obtaining and maintaining the skills required for the procedure. We describe the detailed anatomy of cervical and femoral regions in rats and discuss the advantages for practicing microvascular anastomosis. METHODS: Cervical regions of Sprague-Dawley rats were dissected under intraperitoneal anesthesia. The step-by-step anatomic description was documented using a high-resolution charge-coupled device image sensor and recording systems. Using this model, temporal occlusion time and patency were measured, and these measures were compared between the trainee and trainer groups. The number of times the training needs to be completed to attain competency in the bypass procedure was estimated. RESULTS: After exposing the carotid triangle, a half-ring was created by end-to-side anastomosis. Anastomosis was performed at the common carotid artery using the contralateral side of the carotid artery as a graft. The cutoff value for the temporal occlusion time was 79.3 minutes in the receiver operating characteristic curve based on a target temporal occlusion time for beginners determined during the training. CONCLUSIONS: Using a living animal model, a trainee has the opportunity to learn not only anastomotic techniques but also hemostatic control as well as overcoming mental strain during surgery. Living animal models are important in training because the fidelity of a living animal model is superior to nonliving models. Applying training using a half-ring model contributes to safe and efficient surgery.

Can a Small Intestine Segment Be an Alternative Biological Conduit for Peripheral Nerve Regeneration?

BACKGROUND: Autologous nerve grafts are used to bridge peripheral nerve defects. Limited sources and donor site morbidity are the major problems with peripheral nerve grafts. Although various types of autologous grafts such as arteries, veins and muscles have been recommended, an ideal conduit has not yet been described. AIMS: To investigate the effectiveness of a small intestinal conduit for peripheral nerve defects. STUDY DESIGN: Animal experimentation. METHODS: Twenty-one rats were divided into three groups (n=7). Following anaesthesia, sciatic nerve exploration was performed in the Sham group. The 10 mm nerve gap was bridged with a 15 mm ileal segment in the small intestinal conduit group and the defect was replaced with orthotopic nerve in autologous nerve graft group. The functional recovery was tested monthly by walking-track analysis and the sciatic functional index. Histological evaluation was performed on the 12th week. RESULTS: Sciatic functional index tests are better in autologous nerve graft group (-55.09±6.35); however, during follow-up, progress in sciatic functional index was demonstrated, along with axonal regeneration and innervation of target muscles in the small intestinal conduit group (-76.36±12.08) (p<0.05). In histologic sections, distinctive sciatic nerve regeneration was examined in the small intestinal conduit group. The expression of S-100 and neurofilament was observed in small intestinal conduit group but was less organised than in the autologous nerve graft group. Although the counted number (7459.79±1833.50 vs. 4226.51±1063.06 mm2), measured diameter [2.19 (2.15-2.88) vs. 1.74 (1.50-2.09) µm] and myelin sheath thickness [1.18 (1.09-1.44) vs. 0.66 (0.40-1.07) µm] of axons is significantly high in the middle sections of autologous nerve graft compared to the small intestinal conduit group, respectively (p<0.05), the peripheral nerve regeneration was also observed in the small intestinal conduit group. CONCLUSION: Small intestinal conduit should not be considered as an alternative to autologous nerve grafts in its current form; however, the results are promising. Even though the results are no better than autologous nerve grafts, with additional procedures, it might be a good alternative due to harvesting abundant sources without donor site morbidity.

Efecto de la yerba mate (ilex paraguariensis) sobre el hueso de ratas ovariectomizadas / Effect of yerba mate (ilex paraguariensis) on bone in ovariectomized rats

El consumo de yerba mate (Ilex paraguariensis) es habitual en la Argentina y otros países de América del Sur. La yerba mate, al igual que el café y el té, contiene xantinas y polifenoles. El consumo de café ha mostrado tener impacto negativo sobre la densidad mineral ósea (DMO), mientras que el té ha mostrado tener efecto protector. En mujeres posmenopáusicas tomadoras de mate se halló mayor DMO de columna lumbar y cuello femoral en comparación con controles que no bebían mate. La DMO también fue mayor en ratas que recibieron una infusión de yerba mate y dieta baja en calcio; sin embargo, este incremento no fue capaz de revertir el efecto negativo del bajo contenido de calcio sobre las propiedades biomecánicas y la conectividad trabecular. El objetivo de este trabajo fue estudiar el tejido óseo de ratas ovariectomizadas (OVX) luego de recibir una infusión de yerba mate por 90 días en reemplazo del agua de bebida. Se utilizaron ratas Sprague Dawley (n=16) hembras divididas en 2 grupos: OVX+agua y OVX+yerba. Otro grupo de ratas (n=6) fue sometido a una cirugía simulada (Sham). El estudio mostró claramente el efecto deletéreo de la ovariectomía sobre todos los parámetros estudiados (DMO, histomorfometría ósea, conectividad trabecular y biomecánica) respecto del grupo Sham. El grupo OVX+yerba no mostró diferencias con el grupo OVX+agua en ninguno de los parámetros analizados, por lo que la yerba mate no produciría efecto alguno sobre el hueso de ratas adultas ovariectomizadas. (AU)

Yerba mate (Ilex paraguariensis) consumption is common in Argentina and other South American countries. Like coffee and tea, yerba mate contains xanthines and polyphenols. It has been reported that caffeine consumption has a negative impact on bone mineral density (BMD) while tea has been shown to have a protective effect. On the other hand, in postmenopausal women that usually consumed yerba mate, lumbar spine and femoral neck BMDs were higher than in non-consumers. BMD was also higher in rats that received a yerba mate infusion and low calcium diet. However, this increase was not sufficient to reverse the negative effect of a low calcium diet on bone biomechanical properties and trabecular connectivity. The aim of this work was to study bone tissue in ovariectomized (OVX) rats receiving an infusion of yerba mate instead of drinking water for 90 days. Female Sprague Dawley rats (n=16) were divided into 2 groups: OVX+water and OVX+yerba. A third group of rats (n=6) was submitted to sham surgery (Sham). Results clearly showed the deleterious effect of ovariectomy on all studied parameters (BMD, bone histomorphometry, trabecular connectivity and biomechanical properties) compared to Sham group. The OVX+yerba group showed no difference with OVX+water group in all analyzed parameters. It is concluded that yerba mate does not produce any effect on the bone of ovariectomized adult rats. (AU)

Postoperative pain in Sprague Dawley rats after liver biopsy by laparotomy versus laparoscopy.

Laparoscopic surgery offers advantages for both animal welfare and quality of experimental data. Compared with laparotomy, laparoscopy is associated with less postoperative pain and faster recuperation in humans and is also associated with less postoperative pain in dogs. Postoperative pain associated with laparotomy and laparoscopy has not been compared in rodents, however. The authors used a validated pain grimace scale to evaluate postoperative pain in male Sprague Dawley rats after liver biopsy by laparotomy or laparoscopy. Rats that underwent laparoscopy showed fewer recognized signs of pain than did rats that underwent laparotomy. The authors suggest that laparoscopy could be used for repeated biopsies in rats, minimizing the number of animals used in pharmacological and toxicological studies.

Reducing the number of animals used for microsurgery training programs by using a task-trainer simulator.

To master the skills needed for microsurgery techniques, residents must enrol in a long and complex training program that includes manipulations on simulators, on ex vivo tissues and finally in vivo training. This final step consists of performing vascular anastomoses on murine models. We propose here a simulation program designed to decrease the number of rats used during the final in vivo training. Our study presents the materials used, the various exercises proposed and their evaluations. Two identical student groups were compared in the framework of the University Diploma of Microsurgery. Group A (seven students) followed a classic training program, all of whom achieved permeable vascular anastomoses. A total of 149 rats were needed for this group. Group B (seven students) first validated their manipulations on the task-trainer simulation program. A mean of 6 h was necessary to obtain this validation. All these students achieved the required permeable vascular anastomoses but only 77 rats were used for this group. This simulation program spared 72 rats, abiding by the Russell and Burch concept of a humane experimental technique, namely the 3R principles. This home-made, cost-efficient and easy-to-use task trainer included various exercises with increasing difficulty levels and a progressive scoring system. We believe that microsurgery training needs to include both simple and sophisticated tools in order to reduce the number of animals used to master these surgical skills.

In vitro labeling of human umbilical cord mesenchymal stem cells with superparamagnetic iron oxide nanoparticles.

Human umbilical cord mesenchymal stem cells (hUC-MSCs) transplantation has been shown to promote regeneration and neuroprotection in central nervous system (CNS) injuries and neurodegenerative diseases. To develop this approach into a clinical setting it is important to be able to follow the fates of transplanted cells by noninvasive imaging. Neural precursor cells and hematopoietic stem cells can be efficiently labeled by superparamagnetic iron oxide (SPIO) nanoparticle. The purpose of our study was to prospectively evaluate the influence of SPIO on hUC-MSCs and the feasibility of tracking for hUC-MSCs by noninvasive imaging. In vitro studies demonstrated that magnetic resonance imaging (MRI) can efficiently detect low numbers of SPIO-labeled hUC-MSCs and that the intensity of the signal was proportional to the number of labeled cells. After transplantation into focal areas in adult rat spinal cord transplanted SPIO-labeled hUC-MSCs produced a hypointense signal using T2-weighted MRI in rats that persisted for up to 2 weeks. This study demonstrated the feasibility of noninvasive imaging of transplanted hUC-MSCs.

Efficient processing of laser speckle contrast images.

Though laser speckle contrast imaging enables the measurement of scattering particle dynamics with high temporal resolution, the subsequent processing has previously been much slower. In prior studies, generating a laser speckle contrast image required about 1 s to process a raw image potentially collected in 10 ms or less. In this paper, novel algorithms are described which are demonstrated to convert 291 raw images per second to laser speckle contrast images and as many as 410 laser speckle contrast images per second to relative correlation time images. As long as image processing occurs during image acquisition, these algorithms render processing time irrelevant in most circumstances and enable real-time imaging of blood flow dynamics.

Electrocardiographic changes in rats undergoing thoracic surgery under combined parenteral anesthesia.

To compare two protocols of combined parenteral general anesthesia, the authors analyzed electrocardiographic changes in anesthetized rats undergoing left pneumonectomy. One group of rats was anesthetized with a combination of medetomidine and ketamine (group 1, n = 10), and the other was injected with diazepam and ketamine (group 2, n = 10). Investigators obtained two electrocardiograms from each rat, one before surgery (5 min after anesthesia) and one after surgery (60 min after anesthesia). Anesthetic induction was quick for all rats, though four rats in group 2 died before surgery. Mean cardiac frequency and R-wave amplitude were significantly lower in rats in group 1 than in rats in group 2. Rats in group 1 received injections of atipamezole about 60 min after surgery, which reversed the effects of medetomidine; these rats regained voluntary respiratory movement more quickly than did rats in group 2. Two additional rats in group 2 died during postsurgical recovery. These results suggest that for thoracic surgery in rats, medetomidine-ketamine is an appropriate anesthetic combination, may be safer than diazepam-ketamine and yields a shorter recovery time.

[Effects of estradiol intervention on peripheral blood CD90+ cells and cardiac function in de-ovary rats before and after myocardial infarction].

AIM: To investigate the effects of estradiol intervention on peripheral blood CD90+ cells and cardiac function in de-ovary rats before and after myocardial infarction. METHODS: Thirty SD female rats were divided into the control group, acute myocardial infarction group, de-ovary myocardial infarction group, de-ovary replacement group with estradiol, and de-ovary therapy group with estradiol. CD90+ cell percentage was determined with flow cytometry. The left ventricle end systolic pressure (LVESP), left ventricle end diastolic pressure (LVEDP), +dp/dt(max) and -dp/dt(max) were also measured with the POWERLAB4.12 system. RESULTS: The percentage of peripheral blood CD90+ cells in the de-ovary myocardial infarction group was remarkably lower than that in the acute myocardial infarction group and the de-ovary replacement group with estradiol (P<0.01). In the de-ovary replacement group with estradiol, the percentage of peripheral blood CD90+ cells was remarkably elevated following the first day of operation (P<0.05), reaching the peak value at day 3. In the de-ovary therapy group with estradiol, it started to elevate at day 7 after the operation, much lower than that in the de-ovary replacement group with estradiol. LVESP in each operation group decreased, especially in the acute myocardial infarction group, whereas LVEDP increased in the de-ovary group and de-ovary therapy group with estradiol (P<0.01). CONCLUSION: Replacement with estradiol before myocardial infarction exerts more beneficial effects than estradiol replacement after myocardial infarction in de-ovary rats.

Ketamine-xylazine-acepromazine anesthesia and postoperative recovery in rats.

We evaluated the effect of ketamine-xylazine-acepromazine anesthesia (31.25, 6.25, and 1.25 mg/kg subcutaneously, respectively) on postsurgical recovery in male Sprague-Dawley (Crl:SD) rats undergoing laparotomy with and without the postoperative analgesic ketorolac. Recovery was determined by changes in body weight (BW) and water intake. The time of ketorolac administration (5 mg/kg intramuscularly), 60 min after anesthetic injection, was based on return of the pedal withdrawal reflex in Long-Evans (HsdBlu:LE) rats undergoing stereotaxic surgery in a separate experiment. Results were compared with those of housing and anesthesia controls as well as of laparotomized rats receiving a single sugared treat for nonpharmacologic management of postoperative pain. Surgery took place on day 0; the first 24 h postsurgery was considered the "acute phase," and days 1 through 4 comprised the "recovery phase." Results suggest that 1) the anesthetic mixture is fast- and long-acting and provides sufficient immobility, loss of consciousness, and analgesia; 2) during the acute phase, rats subjected to laparotomy did not lose more BW than rats exposed to anesthesia alone; 3) water intake during both phases did not significantly differ between treatment groups; 4) postsurgical ketorolac administration did not minimize BW loss during the acute phase nor cause any adverse effects under this anesthetic regimen; and 5) provision of single sugared treats had salutary effects on BW recovery. This finding suggests that postsurgical BW loss after use of this anesthetic mixture is due to distress unrelated to pain; this nonpain distress may have masked potential beneficial effects of ketorolac.

Empleo del tisuacryl como hemostático, en hemorragias intrabdominales provocadas en la rata / Use of tisuacryl as an hemostatic in intrabadominal hemorrhages provoked in rats

Se propuso probar los efectos hemostáticos del tisuacryl, biomaterial de producción nacional, en hemorragias provocadas en órganos parenquimatosos de la rata y estudiar histológicamente si ocasiona lesión o no en estos tejidos. La principal indicación de los adhesivos quirúrgicos es el cierre de heridas de piel, pero también se han empleado en urgencias para controlar sangramientos leves en cirugía. Se emplearon 10 ratas, anestesiadas con pentobarbital sódico. Se realizaron heridas en el hígado, bazo y riñón con el bisturí, para provocar una hemorragia moderada y se procedió a hacer la hemostasia con el tisuacryl, hasta cohibir el sangramiento. Los animales se sacrificaron según esquema de evolución diseñado. Se concluyó que el tisuacryl resultó efectivo como hemostático en la rata y desde el punto de vista anatomopatológico, no causó lesión tisular alguna en los tejidos en los cuales se aplicó, por lo que debería utilizarse en la clínica quirúrgica y en los casos de emergencia como arsenal terapéutico(AU)

Systemic bone formation with weekly PTH administration in ovariectomized rats.

PURPOSE: Weekly subcutaneous administration of 0 (vehicle), 10 and 80 microg/kg doses of human parathyroid hormone (1-34) [PTH (1-34)] were compared based on their capacity to induce systemic formation of bone in 9 month-old ovariectomized (OVX) Sprague-Dawley rats. METHODS: Changes elicited at bone tissue after 4 weeks of treatment were assessed using dual x-ray absorptiometry, micro-computed tomography (microCT), and ashing. RESULTS: The 10 microg/kg dose led to a significant increase (p<0.025) in femoral bone mineral density (BMD) over vehicle- and 80 microg/kg-treated groups. Similarly, structural analysis of the femoral neck trabecular bone by microCT revealed increases in bone volume fraction and trabecular thickness over the pre-treatment baseline, and vehicle- and 80 microg/kg-treated groups. CONCLUSIONS: The data suggest that the weekly administration of 10 microg/kg of PTH (1-34) was sufficient to significantly promote the bone mineral density systemically. The weekly administration of 10 microg/kg over a 4-week treatment period is, to our knowledge, one of the lowest reported total dose of PTH (1-34) shown to induce a net anabolic effect on skeletal tissue in OVX rats.

A stepwise surgical procedure to investigate the lymphatic transport of lipid-based oral drug formulations: Cannulation of the mesenteric and thoracic lymph ducts within the rat.

INTRODUCTION: A number of animal models have been described for the assessment of intestinal lymphatic drug transport. Lymphatic transport studies are commonly first conducted in the laboratory rat, with larger more complicated models (i.e., dog or pig) subsequently investigated. However, the utility of lymph fistulation in large animals is limited by considerable logistical and economic constraints. METHODS: This paper describes a stepwise surgical procedure for cannulating the thoracic and mesenteric lymph ducts in male Sprague-Dawley rats. RESULTS: Following surgery, thoracic and mesenteric lymph flow rates during the 24-h period immediately following surgery averaged 12.5+/-2.5 and 2.4+/-1.1 ml/h, respectively. This flow rate is greater than that obtained with previously described methods, which require restraint of the animals and/or a 24-h recovery period and are reported to produce average intestinal lymph flow rates of 2 ml/h. DISCUSSION: This animal model can be utilized for the assessment of drug transport by the lymphatics and for determining what percentage of lymphatic transport is a result of only intestinal lymphatics.

Continuous administration of the 5-hydroxytryptamine1A agonist (3-Chloro-4-fluoro-phenyl)-[4-fluoro-4-[[(5-methyl-pyridin-2-ylmethyl) -amino]-methyl]piperidin-1-yl]-methadone (F 13640) attenuates allodynia-like behavior in a rat model of trigeminal neuropathic pain.

(3-Chloro-4-fluoro-phenyl)-[4-fluoro-4-[[(5-methyl-pyridin-2-ylmethyl)-amino]-methyl]piperidin-1-yl]-methadone (F 13640) is a recently discovered high-efficacy 5-hydroxytryptamine (HT)1A receptor agonist that produces central analgesia through the neuroadaptive mechanisms of inverse tolerance and cooperation. In a rat model of trigeminal neuropathic pain, the chronic constriction injury of the infraorbital nerve causes allodynia-like behavior that develops within 2 weeks and remains stable thereafter. We report that early after surgery, during which time allodynia develops, the continuous 2-week infusion of 0.63 mg/day F 13640 inhibited the allodynia-like behavior, whereas 5 mg/day morphine showed no significant effect. When F 13640 infusion was initiated late after surgery, when allodynia was well established, it produced an antiallodynic effect that was apparent during the entire infusion period. In contrast, morphine infusion caused an initially marked antiallodynic effect to which tolerance developed within the 2-week infusion period. The GABA-B receptor agonist baclofen (1.06 mg/day) that has a recognized usefulness in the treatment of trigeminal neuralgia, demonstrated effectiveness in both conditions. The data are consistent with a theory of nociceptive signal transduction, as well as with previous data, in demonstrating the neuroadaptive mechanisms of inverse tolerance and cooperation. That is, in contrast with morphine, the antiallodynic effect induced by 5-HT1A receptor activation does not decay, but, if anything, grows with chronicity. Also, 5-HT1A receptor activation seemed to cooperate with nociceptive stimulation in, paradoxically, inducing an antiallodynic effect. The data presented here suggest that F 13640 may perhaps offer a lasting treatment of trigeminal neuralgia.

Estudio experimental de homoinjertos criopreservados: modificaciones miointimales

Objetivo. Evaluar el desarrollo de hiperplasia intimal, muerte celular y respuesta macrofágica, en injertos arteriales criopreservados tras su implante en animales de experimentación. Material y métodos. Arterias ilíacas de rata Spraque-Dawley fueron criopreservadas de manera automatizada, en un congelador biológico, con un rango de disminución de temperatura de 1ºC/min. Fueron almacenadas a 145ºC en vapores de N2 líquido durante 30 días y sometidas a descongelación lenta y computerizada. Los injertos singénicos, se efectuaron a nivel de la arteria ilíaca común. Se establecieron los siguientes grupos de estudio: GI: arterias criopreservadas no implantadas, GII: microinjertos frescos implantados y GIII: arterias criopreservadas e implantadas. Manteniendo como grupo control (GC), arterias ilíacas frescas. Los animales fueron sacrificados a los 14, 30 y 90 días post-injerto. Se valora la hiperplasia intimal sobre los injertos mediante morfometría, se realizan técnicas de TUNEL y respuesta macrofágica. Resultados. La permeabilidad en el momento del sacrificio fue del 100 per cent para homoinjertos fresco y 66.6 per cent para los implantes criopreservados. El propio proceso de criopreservación (GI) indujo daño en la pared arterial sobre todo a nivel endotelial mostrando áreas de denudación de diferente extensión y una buena celularidad de la capa media. La hiperplasia intimal sobre el injerto a los 14 días de implante (GII), estaba visiblemente retrasada hasta los 30 días cuando el injerto era criopreservado (GIII). Las arterias de este último grupo, mostraban un adelgazamiento general de la pared arterial y degeneración con pérdida de celularidad a nivel de la capa media del injerto. La presencia macrofágica fue similar en los dos grupos implantados, limitándose a zonas de anastomosis e hiperplasia. Los estudios de daño celular mostraron mayor número de células marcadas en el GIII a los 90 días post-implante. Conclusiones. La respuesta hiperplásica se modifica cuando el injerto ha sido previamente criopreservado, el retraso en su formación y una mayor tasa de muerte celular a largo plazo, sin alteraciones en la respuesta macrofágica, parecen ser indicativos de una clara tendencia a la degeneración del mismo a largo plazo (AU)

Electroporation for direct spinal gene transfer in rats.

We investigated the feasibility of delivering exogenous genes into spinal cord using direct in vivo electrotransfection. Gene transfer to the spinal cord was accomplished via direct intrathecal injection of pE-GFP C1 vector, followed by five electric pulses for 50 ms at 200 V delivered intrathecally. The spinal cords were retrieved and analyzed with fluorescence microscopy, reverse transcription polymerase chain reaction (RT-PCR), and Western blotting. At day 1, 3 or 7 following electroporation a clear GFP expression in spinal cord tissue was detected. The most prominent transfection occurred in the meningeal cells and superficial layer of the spinal cord. Successful transfection was also confirmed with RT-PCR and Western blotting. The expression of GFP protein was peaked between 3 and 7 days after electroporation and significantly decreased at 14 days. No behavioral or spinal neurodegenerative changes were detected at any time point. This study demonstrates that direct in vivo electrotransfection represents an effective and simple method for spinal gene delivery and have a potential to be used clinically, especially, acute or chronic pain.