Inadvertent injection of medetomidine in the cerebromedullaris cisterna of a dog during myelographic exam.

INTRODUCTION: A mixed breed dog was anesthetized for diagnostic myelography to investigate acute onset neck pain. Instead of contrast medium, 444 µg/kg medetomidine were inadvertently injected into the cerebromedullaris cisterna owing to a human error. Severe bradycardia, undetectable peripheral pulse, respiratory arrest and loss of pupillary, palpebral and corneal reflexes were observed immediately after injection. Profound hypothermia developed and esophageal temperature, measured 20 minutes after medetomidine injection, was 33 °C. Atipamezole at 1 mg/kg im was administered, followed by a second dose of 0,5 mg/kg iv 20 minutes thereafter. In the meantime, cardiorespiratory parameters and body temperature were monitored, and supportive care that included manually assisted pulmonary ventilation, active warming, and administration of 5 µg/kg/min dopamine was initiated. The dog's clinical condition improved within one hour from the beginning of supportive care, at which time ocular reflexes and swallowing returned, spontaneous ventilation was deemed as adequate and the trachea could be extubated. The dog was discharged in good clinical conditions five days later. Human error and distraction led to a potentially life-threatening complication in the dog of this report and could have possibly been prevented with the use of checklists and with a clearer definition of roles and responsibilities of the personnel involved prior to commencing the clinical procedure. Profound cardiovascular, respiratory, and thermoregulatory depression caused by intracisternal injection of medetomidine responded to parenteral administration of its antagonist and supportive care.

INTRODUCTION: Un chien croisé a été anesthésié pour une myélographie diagnostique afin d'étudier une douleur aiguë au niveau du cou. Au lieu du produit de contraste, 444 µg/kg de médétomidine ont été injectés par inadvertance dans la citerne cérébello-médullaire en raison d'une erreur humaine. Une bradycardie sévère, un pouls périphérique indétectable, un arrêt respiratoire et une perte des réflexes pupillaire, palpébral et cornéen ont été observés immédiatement après l'injection. Une hypothermie profonde s'est développée et la température oesophagienne, mesurée 20 minutes après l'injection de médétomidine, était de 33 °C. De l'atipamézole à 1 mg/kg im a été administré, suivi d'une seconde dose de 0,5 mg / kg iv 20 minutes après. Dans l'intervalle, les paramètres cardiorespiratoires et la température corporelle ont été surveillés et des soins de soutien comprenant une ventilation assistée manuellement, un réchauffement actif et l'administration de 5 µg/kg/min de dopamine ont été initiés. L'état clinique du chien s'est amélioré dans l'heure qui a suivi le début des soins, moment où les réflexes oculaires et la déglutition sont réapparus, la ventilation spontanée a été jugée adéquate et où on a pu procéder à l'extubation. Le chien est sorti dans de bonnes conditions cliniques cinq jours plus tard. Une erreur humaine et de la distraction ont conduit à une complication potentiellement mortelle chez le chien décrit dans ce rapport et auraient pu être évitées grâce à l'utilisation de listes de contrôle et avec une définition plus claire des rôles et des responsabilités du personnel impliqué avant le début de la procédure clinique. Une profonde dépression cardiovasculaire, respiratoire et de la thermorégulation causée par l'injection intracisternale de médétomidine a répondu à l'administration parentérale de son antagoniste et à des soins de soutien.

Impediment of Cerebrospinal Fluid Drainage Through Glymphatic System in Glioma.

BACKGROUND: Cerebrospinal fluid (CSF) plays an important role in maintaining tissue homeostasis in the central nervous system. In 2012, the new CSF outflow pathway, "the glymphatic system," was discovered. The glymphatic system mediates CSF and interstitial fluid exchange through the perivascular pathway, which eliminates harmful solutes in the brain parenchyma. In recent studies, the importance of the glymphatic system has been demonstrated in healthy and neurodegenerative disease brains. However, there is limited research on the function of the CSF in brain tumors. Intracranial hypertension caused by glioma can affect CSF drainage, which impacts the delivery of chemotherapy drugs via intrathecal injection. This study focused on changes in the glymphatic system and the role of aquaporin 4 (AQP4) in glymphatic transport in glioma. METHODS: In glioma-bearing rats, the effect of tracer infusion on the intracranial pressure (ICP) was evaluated using an ICP microsensor. In vivo magnetic resonance imaging and ex vivo bright field were used to monitor CSF tracer distribution after cisterna magna injection. AQP4 expression was quantitatively detected, and AQP4 in the astrocytes around the vessels was observed using immunofluorescence. RESULTS: The ICP of the tumor group was higher than that of the control group and the infusion rate of 2 µl/min did not affect ICP. In vivo and ex vivo imaging showed that the circulation of CSF tracers was significantly impaired in the tumor. High-power confocal microscopy revealed that, in the tumor, the surrounding of AQP4 by Evans Blue was decreased. In both tumor and contralateral areas, data indicated that the number of cluster designation 34 (CD34+) alpha-smooth muscle actin (α-SMA-) veins were more than that of CD34+α-SMA+ arteries. Moreover, in the tumor area, AQP4 in the astrocytes around the vessels was decreased. CONCLUSIONS: These findings indicate that the para-arterial influx of subarachnoid CSF is limited in glioma, especially in those with reduced levels of the fundamental protein AQP4. Our results provide evidence toward a potential new treatment method for glioma in the future.

Gene Therapy in a Mouse Model of Niemann-Pick Disease Type C1.

Niemann-Pick disease type C1 (NPC1) is a fatal congenital neurodegenerative disorder caused by mutations in the NPC1 gene, which is involved in cholesterol transport in lysosomes. Broad clinical manifestations of NPC1 include liver failure, pulmonary disorder, neurological deficits, and psychiatric symptoms. The main cause of death in NPC1 patients involves central nervous system (CNS) dysfunction; there is no essential treatment. We generated a tyrosine-mutant adeno-associated virus (AAV) 9/3 vector that expresses human NPC1 under a cytomegalovirus (CMV) promoter (AAV-CMV-hNPC1) and injected it into the left lateral ventricle (5 µL) and cisterna magna (10 µL) of Npc1 homo-knockout (Npc1-/-) mice. Each mouse received total 1.35 × 1011 vector genome on days 4 or 5 of life. AAV-treated Npc1-/- mice (n = 11) had an average survival of >28 weeks, while all saline-treated Npc1-/- mice (n = 11) and untreated Npc1-/- mice (n = 6) died within 16 weeks. Saline-treated and untreated Npc1-/- mice lost body weight from 7 weeks until death. However, the average body weight of AAV-treated Npc1-/- mice increased until 15 weeks. AAV-treated Npc1-/- mice also showed a significant improvement in the rotarod test performance. A pathological analysis at 11 weeks showed that cerebellar Purkinje cells were preserved in AAV-treated Npc1-/- mice. In contrast, untreated Npc1-/- mice showed an almost total loss of cerebellar Purkinje cells. Combined injection into both the lateral ventricle and cisterna magna achieved broader delivery of the vector to the CNS, leading to better outcomes than noted in previous reports, with injection into the lateral ventricles or veins alone. In AAV-treated Npc1-/- mice, vector genome DNA was detected widely in the CNS and liver. Human NPC1 RNA was detected in the brain, liver, lung, and heart. Accumulated unesterified cholesterol in the liver was reduced in the AAV-treated Npc1-/- mice. Our results suggest the feasibility of gene therapy for patients with NPC1.

Rat Cerebrospinal Fluid Treatment Method through Cisterna Cerebellomedullaris Injection.

Drugs that lack the ability to cross the blood-brain barrier (BBB) need to be placed directly into the central nervous system. Our laboratory studies the involvement of the glutamatergic system in the aggressiveness of glioma, and some ligands of glutamate receptors cannot permeate the BBB. Here, glioma-implanted rats were treated by a technique that delivers ligands directly into the cerebrospinal fluid by puncture into the cisterna cerebellomedullaris. Rats were anesthetized and fixed in a rodent stereotactic device. The head was gently tilted downwards at an angle that allowed exposure of the cisterna. Injection into the cisterna was done freehand using a gingival needle coupled to a microsyringe. The efficiency of intracisternal injection was demonstrated using a methylene blue solution. This type of injection is adaptable for any rodent model using small volumes of a variety of other drugs, and is an interesting method for neuroscience studies.

Establishment of a rat model of Cryptococcus neoformans meningitis / 中国感染与化疗杂志

Objective To establish a rat model of Cryptococcus neoformans meningitis. Methods Sprague-Dawley rats were divided into four groups (group A to D). The load of Cryptococcus neoformans was inoculated by intracisternal injection to the animals in Group A (1×105 cfu), Group B (1×106 cfu), Group C (1×107 cfu), and 0.9％ NaCl solution to Group D. The rats were observed after inoculation for their clinical symptoms. On day 14 and day 21 after inoculation, cerebrospinal fluid was sampled and cultured for counting of bacterial colonies. The 28-day mortality of the animals was calculated. Results All the animals in group A (1×105 cfu) survived without any apparent clinical symptoms, and associated with decreasing bacteria load. The animals in group B (1×106 cfu) had mild symptoms associated with low mortality rate and slightly increased bacterial load. The animals in group C (1×107 cfu) showed a lot of symptoms and associated with high mortality rate and significantly increased bacteria load. All the animals in group D (0.9％ NaCl solution) survived with normal activity. No bacterial colony was cultured from the cerebrospinal fluid. Conclusions Intracisternal injection of 1×107 cfu Cryptococcus neoformans to rats could cause apparent clinical symptoms of meningitis. The 28-day mortality rate of such a rat model of Cryptococcus neoformans meningitis is greater than 80％. An ideal rat model of Cryptococcus neoformans meningitis is established successfully.

Rat Cerebrospinal Fluid Treatment Method through Cisterna Cerebellomedullaris Injection / 神经科学通报·英文版

Drugs that lack the ability to cross the blood-brain barrier (BBB) need to be placed directly into the central nervous system. Our laboratory studies the involvement of the glutamatergic system in the aggressiveness of glioma, and some ligands of glutamate receptors cannot permeate the BBB. Here, glioma-implanted rats were treated by a technique that delivers ligands directly into the cerebrospinal fluid by puncture into the cisterna cerebellomedullaris. Rats were anesthetized and fixed in a rodent stereotactic device. The head was gently tilted downwards at an angle that allowed exposure of the cisterna. Injection into the cisterna was done freehand using a gingival needle coupled to a microsyringe. The efficiency of intracisternal injection was demonstrated using a methylene blue solution. This type of injection is adaptable for any rodent model using small volumes of a variety of other drugs, and is an interesting method for neuroscience studies.

In vivo bioluminescence imaging for leptomeningeal dissemination of medulloblastoma in mouse models.

BACKGROUND: The primary cause of treatment failure in medulloblastomas (MB) is the development of leptomeningeal dissemination (seeding). For translational research on MB seeding, one of the major challenges is the development of reliable experimental models that simulate the seeding and growth characteristics of MBs. To overcome this obstacle, we improved an experimental mouse model by intracisternal inoculation of human MB cells and monitoring with in vivo live images. METHODS: Human MB cells (UW426, D283 and MED8A) were transfected with a firefly luciferase gene and a Thy1.1 (CD90.1) marker linked with IRES under the control of the CMV promoter in a retroviral DNA backbone (effLuc). The MB-effLuc cells were injected into the cisterna magna using an intrathecal catheter, and bioluminescence images were captured. We performed histopathological analysis to confirm the extent of tumor seeding. RESULTS: The luciferase activity of MB-effLuc cells displayed a gradually increasing pattern, which correlated with a quantitative luminometric assay. Live imaging showed that the MB-effLuc cells were diffusely distributed in the cervical spinal cord and the lumbosacral area. All mice injected with UW426-effLuc, D283-effLuc and MED8A-effLuc died within 51 days. The median survival was 22, 41 and 12 days after injection of 1.2 × 10(6) UW426-effLuc, D283-effLuc and MED8A-effLuc cells, respectively. The histopathological studies revealed that the MB-effLuc cells spread extensively and diffusely along the leptomeninges of the brain and spinal cord, forming tumor cell-coated layers. The tumor cells in the subarachnoid space expressed a human nuclei marker and Ki-67. Compared with the intracerebellar injection method in which the subfrontal area and distal spinal cord were spared by tumor cell seeding in some mice, the intracisternal injection model more closely resembled the widespread leptomeningeal seeding observed in MB patients. CONCLUSION: The results and described method are valuable resources for further translational research to overcome MB seeding.

Effect of Endothelin-1 on Baroreflexes and the Cardiovascular Action of Clonidine in Conscious Rabbits.

We studied the influence of pretreatment with endothelin-1 on cardiac baroreflexes and on the effect of clonidine on blood pressure and heart rate. In order to avoid the complication of the direct vasoconstrictor effects of endothelin-1, initial dose-response studies in animals treated with a ganglion blocker were performed. Intravenous administration of 50, 200, and 1200 ng/kg of endothelin-1 produced biphasic changes in blood pressure, consisting of an immediate depressor response, followed by a long lasting and dose-dependent pressor effect (peak response 3 ± 1, 9 ± 3, and 33 ± 5 mmHg, respectively). Thus, the 50 ng/kg dose of endothelin-1 was used in subsequent studies. Conscious rabbits were pretreated on separate days with endothelin-1, either intravenously (50 ng/kg) or intracisternally (10 and 50 ng/kg), or with vehicle. The animals then received an intravenous dose (20 µg/kg) or an intracisternal dose (1 µg/kg) of clonidine and the effects on blood pressure and heart rate were measured. In vehicle-treated rabbits, the intravenous administration of clonidine induced a significant decrease in blood pressure and heart rate (15 min after injection: -15.7 ± 4.7 mmHg and -33 ± 4 b/min, respectively). Similarly, the intracisternal injection of clonidine lowered blood pressure (-16.0 ± 2.5 mmHg), but produced a less pronounced bradycardia (-18 ± 4 b/min). Endothelin pretreatment, either 50 ng/kg centrally or peripherally, had no significant effect on the hypotension or bradycardia produced either by central or peripheral injection of clonidine. At this dose, endothelin by itself did not produce significant changes in blood pressure or heart rate. There was a reduction of the gain of the baroreceptor-heart rate reflex with intracisternal endothelin-1. These results suggest that central 2-adrenoceptor mechanisms involved in clonidine-induced hypotension and bradycardia do not appear to be influenced by activation of endothelin receptors.

Itch elicited by intradermal injection of serotonin, intracisternal injection of morphine, and their synergistic interactions in rats.

We used the cheek model of itch and pain in rats to determine the dose-response relationships for intradermal injection of serotonin and α methylserotonin on scratching behavior. We also determined the dose-related effects of intracisternally injected morphine on scratching, effects that were greatly reduced by administration of the opiate antagonist naloxone. We then examined the interactions of intradermal injection of serotonin and intracisternal injection of morphine on scratching and found that the two procedures act synergistically to increase itch. These results suggest that morphine applied to the CNS is capable of producing itch and greatly increasing itch originating in the skin (hyperknesis).

Protective role of noradrenaline in benzo[a]pyrene-induced learning impairment in developing rat.

Benzo[a]pyrene (B[a]P), a carcinogen, affects brain development, learning, and memory. Isolated studies have reported that B[a]P elevates noradrenaline (NA) level that may modulate neuronal growth, learning, and memory. Therefore, we investigated in vivo and in vitro the effects of B[a]P on learning and memory and its possible mechanism of action. Intracisternal administration of B[a]P on postnatal day 5 significantly reduced learning and memory in adolescent rats as observed by probe test using the Morris water maze. The density of both the subunits of the N-methyl-D-aspartate (NMDA) receptor, NMDAR1 and NMDAR2B, significantly increased in the hippocampus. In vitro, B[a]P significantly increased NMDAR1 in both C6 and Neuro2a cell lines, whereas NMDAR2B was significantly increased in C6 but was significantly decreased in Neuro2a. Pretreatment with NA prevented the B[a]P-induced effect on NMDAR1 expression in both cell lines. However, although NA prevented the B[a]P-mediated increase in NMDAR2B expression in C6, it further potentiated the decrease of NMDAR2B in Neuro2a cells. Also, NA prevented the B[a]P-induced increase in intracellular Ca(2+) both in C6 and in Neuro2a. Our findings show that postnatal exposure of developing rats to B[a]P impairs learning and memory even when the rats became adolescent. We also observed that the effects were mediated by elevated intracellular Ca(2+) levels and increased expression of NMDAR; furthermore, NA exerted a protective effect by modulating those factors. NA differentially affects neurons and glia, which may have a compensatory role during toxic insults, especially from B[a]P.

Food-intake dysregulation in type 2 diabetic Goto-Kakizaki rats: hypothesized role of dysfunctional brainstem thyrotropin-releasing hormone and impaired vagal output.

Thyrotropin-releasing hormone (TRH), a neuropeptide contained in neural terminals innervating brainstem vagal motor neurons, enhances vagal outflow to modify multisystemic visceral functions and food intake. Type 2 diabetes (T2D) and obesity are accompanied by impaired vagal functioning. We examined the possibility that impaired brainstem TRH action may contribute to the vagal dysregulation of food intake in Goto-Kakizaki (GK) rats, a T2D model with hyperglycemia and impaired central vagal activation by TRH. Food intake induced by intracisternal injection of TRH analog was reduced significantly by 50% in GK rats, compared to Wistar rats. Similarly, natural food intake in the dark phase or food intake after an overnight fast was reduced by 56-81% in GK rats. Fasting (48h) and refeeding (2h)-associated changes in serum ghrelin, insulin, peptide YY, pancreatic polypeptide and leptin, and the concomitant changes in orexigenic or anorexigenic peptide expression in the brainstem and hypothalamus, all apparent in Wistar rats, were absent or markedly reduced in GK rats, with hormone release stimulated by vagal activation, such as ghrelin and pancreatic polypeptide, decreased substantially. Fasting-induced Fos expression accompanying endogenous brainstem TRH action decreased by 66% and 91%, respectively, in the nucleus tractus solitarius (NTS) and the dorsal motor nucleus of the vagus (DMV) in GK rats, compared to Wistar rats. Refeeding abolished fasting-induced Fos-expression in the NTS, while that in the DMV remained in Wistar but not GK rats. These findings indicate that dysfunctional brainstem TRH-elicited vagal impairment contributes to the disturbed food intake in T2D GK rats, and may provide a pathophysiological mechanism which prevents further weight gain in T2D and obesity.