Free-Hand Intracerebroventricular Injections in Mice.

The investigation of neuroendocrine systems often requires the delivery of drugs, viruses, or other experimental agents directly into the brains of mice. An intracerebroventricular (ICV) injection allows the widespread delivery of the experimental agent throughout the brain (particularly in the structures near the ventricles). Here, methods for making free-hand ICV injections in adult mice are described. By using visual and tactile landmarks on the heads of mice, injections into the lateral ventricles can be made rapidly and reliably. The injections are made with a glass syringe held in the experimenter's hand and placed at approximate distances from the landmarks. Thus, this technique does not require a stereotaxic frame. Furthermore, this technique requires only brief isoflurane anesthesia, which permits the subsequent assessment of mouse behavior and/or physiology in awake, freely behaving mice. Free-hand ICV injection is a powerful tool for the efficient delivery of experimental agents into the brains of living mice and can be combined with other techniques such as frequent blood sampling, neural circuit manipulation, or in vivo recording to investigate neuroendocrine processes.

Aquaporin 4 Mediates the Effect of Iron Overload on Hydrocephalus After Intraventricular Hemorrhage.

BACKGROUND: Iron overload plays an important role in hydrocephalus development following intraventricular hemorrhage (IVH). Aquaporin 4 (AQP4) participates in the balance of cerebrospinal fluid secretion and absorption. The current study investigated the role of AQP4 in the formation of hydrocephalus caused by iron overload after IVH. METHODS: There were three parts to this study. First, Sprague-Dawley rats received an intraventricular injection of 100 µl autologous blood or saline control. Second, rats had IVH and were treated with deferoxamine (DFX), an iron chelator, or vehicle. Third, rats had IVH and were treated with 2-(nicotinamide)-1,3,4-thiadiazole (TGN-020), a specific AQP4 inhibitor, or vehicle. Rats underwent T2-weighted and T2* gradient-echo magnetic resonance imaging to assess lateral ventricular volume and intraventricular iron deposition at 7, 14, and 28 days after intraventricular injection and were then euthanized. Real-time quantitative polymerase chain reaction, western blot analysis, and immunofluorescence analyses were conducted on the rat brains to evaluate the expression of AQP4 at different time points. Hematoxylin and eosin-stained brain sections were obtained to assess the ventricular wall damage on day 28. RESULTS: Intraventricular injection of autologous blood caused a significant ventricular dilatation, iron deposition, and ventricular wall damage. There was increased AQP4 mRNA and protein expression in the periventricular tissue in IVH rats through day 7 to day 28. The DFX treatment group had a lower lateral ventricular volume and less intraventricular iron deposition and ventricular wall damage than the vehicle-treated group after IVH. The expression of AQP4 protein in periventricular tissue was also inhibited by DFX on days 14 and 28 after IVH. The use of TGN-020 attenuated hydrocephalus development after IVH and inhibited the expression of AQP4 protein in the periventricular tissue between day 14 and day 28 without a significant effect on intraventricular iron deposition or ventricular wall damage. CONCLUSIONS: AQP4 located in the periventricular area mediated the effect of iron overload on hydrocephalus after IVH.

Orexin deficiency modulates the dipsogenic effects of angiotensin II in a sex-dependent manner.

The orexin (hypocretin) neuropeptide system is an important regulator of ingestive behaviors, i.e., it promotes food and water intake. Here, we investigated the role of orexin in drinking induced by the potent dipsogen angiotensin II (ANG II). Specifically, male and female orexin-deficient mice received intracerebroventricular (ICV) injections of ANG II, followed by measuring their water intake within 15 min. We found that lower doses of ANG II (100 ng) significantly stimulated drinking in males but not in females, indicating a general sex-dependent effect that was not affected by orexin deficiency. However, higher doses of ANG II (500 ng) were sufficient to induce drinking in female wild-type mice, while female orexin-deficient mice still did not respond to the dipsogenic properties of ANG II. In conclusion, these results suggest sex-dependent effects in ANG II-induced drinking and further support the sexual dimorphism of orexin system functions.

Role of central adiponectin and its interactions with NPY and GABAergic systems on food intake in neonatal layer chicken.

BACKGROUND & AIM: Adiponectin is a member of the adipokine family and contributes to regulating energy homeostasis, reproduction, and various biological functions, such as insulin receptor signaling pathway sensitivity, mitochondrial biogenesis, oxidative metabolism, neurogenesis, and suppression of inflammation. This study aimed to investigate the effects of intracerebroventricular (ICV) injection of adiponectin and its interaction with the neuropeptide Y (NPY) and GABAergic systems on central appetite regulation in neonatal layer-type chickens. MATERIALS & METHODS: In this study, 6 experiments were conducted, each of which included 4 experimental groups. In the first experiment, the chickens were injected with saline and adiponectin (20.73, 41.45, and 62.18 nmol). In the second experiment, saline, adiponectin (62.18 nmol), B5063 (NPY1 receptor antagonist, 2.12 nmol), and simultaneous injections of adiponectin and B5063 were performed. Experiments 3 to 6 were done in the same way to experiment 1, but the chickens were injected with SF22 (NPY2 receptor antagonist, 2.66 nmol), SML0891 (NPY5 receptor antagonist, 2.89 nmol), picrotoxin (GABAA receptor antagonist, 0.89 nmol), CGP54626 (GABAB receptor antagonist, 0.047 nmol) instead of B5063. Feed consumption was measured 120 min after the injection. RESULTS: A dose-dependent increase in appetite was observed after the injection of adiponectin (20.73, 41.45, and 62.18 nmol) (P < 0.05). The injection of B5063 + adiponectin attenuated the hyperphagic effect of adiponectin (P < 0.05). In addition, co-injection of picrotoxin and adiponectin significantly decreased adiponectin-induced hyperphagia (P < 0.05). In addition, adiponectin significantly increased the number of steps, jumps, exploratory food, pecks, and standing time, while decreasing sitting time and rest time (P < 0.05). CONCLUSION: These results suggest that the hyperphagic effects of adiponectin are probably mediated through NPY1 and GABAA receptors in neonatal layer-type chickens.

Clinical Efficacy of Intraventricular rt-PA (Actilyse®) in the Outcome of Patients with Spontaneous Intraventricular Hemorrhage.

AIM: To evaluate the effectiveness of intraventricular injection of rt-PA (Actilyse®) in patients with spontaneous intraventricular hemorrhage (IVH) who had undergone external ventricular drainage (EVD). MATERIAL AND METHODS: This randomized clinical trial recruited 60 patients with spontaneous IVH who had undergone EVD due to the signs of hydrocephalus. The patients were randomly divided into two groups, including a group receiving intraventricular injection of rt-PA and the other normal saline. RESULTS: Both groups receiving rt-PA Actilyse® (n=28) or placebo (n=32) were male by majority (58.33%). We found no difference in the prevalence of meningitis and brain infection (35.7% vs. 37.5%, p=0.665). Changes in hematoma volume at the end of the fourth day compared to the first day after EVD differed significantly between the two groups (p=0.004). The majority (64.29%) showed a decrease in the rt-PA group, but in the placebo group, the majority (53.13%) remained constant. As a result, changes in the rt-PA group were significantly higher than those in the placebo group. Improvements in the level of consciousness (GCS) at the end of the fourth day compared to the first day after EVD implantation was 1.07 units in the Actilyse® group and -1.91 in the placebo group. As shown, the fourth day showed significant differences between the two groups (p < 0.001). Improvements in the Glasgow Coma Scale (GCS) were observed at the end of the period. CONCLUSION: It can be concluded that intraventricular injection of rt-PA (Actilyse®) can effectively reduce the volume of hematoma and improve the level of consciousness (GCS) during treatment. Intraventricular injection of 2-mg rt-PA is safe for patients and does not cause any acute complications such as cerebral hematoma expansion.

Outcomes of Combined Endoscopic Surgery and Fibrinolytic Treatment Protocol for Intraventricular Hemorrhage: A Randomized Controlled Trial.

BACKGROUND: Intraventricular fibrinolysis (IVF) and endoscopic surgery (ES) are the new promising treatment strategies to enhance the rate of hematoma clearance, which might improve functional outcome. This study investigated and compared the outcomes among these interventions. METHODS: A randomized (1:1) double-blinded trial was carried out between August 2018 and December 2021. The intervention and control groups comprised patients receiving IVF and/or ES and external ventricular drainage (EVD), respectively. All participants had experienced primary or secondary intraventricular hemorrhage (IVH) from spontaneous intracerebral hemorrhage with obstructive hydrocephalus complications. The primary outcome was modified Rankin Scale score 180 days post treatment. Interim assessments were planned for every 50 participants enrolled to ensure safety and efficacy. RESULTS: After enrollment of 110 participants (55 participants in each group), there was a difference in 30-day mortality (2 [3.6%] vs. 13 [32.7%] in the EVD group, P = 0.002), reaching the predetermined boundaries for termination of the trial. We demonstrated a better favorable outcome (modified Rankin Scale score 0-3) at 180 days in the intervention group, compared with the control group (35 [63.6%] vs. 24 [43.6%], P = 0.04). Participants in the intervention group experienced a higher IVH removal rate (91% [9.0] vs. 69.5% [38.0], P < 0.01) and had lower shunt conversion (1 [1.8%] vs. 16 [29.3%], P < 0.01). Treatment complications were comparable between the two groups. CONCLUSIONS: This study demonstrated that combined ES and IVF is safe and effective for the treatment of IVH. In addition, it concluded that aggressive but safe procedures used to remove IVH could improve clinical outcome in patients with IVH.

Meningitis por Listeria: administración de gentamicina intraventricular y monitorización de concentraciones en líquido cefalorraquídeo / Listeria meningitis: administration of intraventricular gentamicin and monitoring of concentrations in cerebrospinal fluid

La Listeria continúa siendo una posible etiología de meningitis bacteriana en nuestro medio, siendo causa más frecuente en neonatos, ancianos o pacientes inmunodeprimidos. Debido a la gravedad y la mortalidad asociada, resulta de gran interés disponer de nuevas herramientas que permitan un manejo clínico y farmacológico más eficaz.Presentamos un caso de meningitis por Listeria que ingresa en la Unidad de Cuidados Intensivos. Dada la escasa penetración de la gentamicina en el sistema nervioso central y siendo ésta uno de los tratamientos de elección en las guías clínicas de referencia, se decide la administración de gentamicina intraventricular llevando a cabo una monitorización de concentraciones de gentamicina en líquido cefalorraquídeo (LCR).Debido a la alta variabilidad farmacocinética del paciente crítico, la monitorización de concentraciones en LCR de gentamicina tras su administración intraventricular puede resultar de gran utilidad para asegurar el alcance de concentraciones de fármaco que permitan una mayor eficacia del tratamiento. (AU)

Listeria is currently a possible etiology of bacterial meningitis in our society, being one more frequent cause in neonates, elderly or immunosuppressed patients. Due to the severity and mortality associated, it is therefore very useful to have new tools that allow a more effective clinical and pharmacological management.We present a case of Listeria meningitis admitted to the Intensive Care Unit. Given the low penetration of gentamicin into the central nervous system and being one of the treatments of choice in the clinical reference guidelines, the administration of intraventricular gentamicin was decided by monitoring the concentrations of gentamicin in cerebrospinal fluid (CSF).Due to the high pharmacokinetic variability of the critically ill patient, monitoring CSF concentrations of gentamicin after intraventricular administration can be very useful to ensure the achievement of drug concentrations that allow greater treatment efficacy. (AU)

Modeling Neonatal Intraventricular Hemorrhage through Intraventricular Injection of Hemoglobin.

Neonatal intraventricular hemorrhage (IVH) is a common consequence of premature birth and leads to brain injury, posthemorrhagic hydrocephalus (PHH), and lifelong neurological deficits. While PHH can be treated by temporary and permanent cerebrospinal fluid (CSF) diversion procedures (ventricular reservoir and ventriculoperitoneal shunt, respectively), there are no pharmacological strategies to prevent or treat IVH-induced brain injury and hydrocephalus. Animal models are needed to better understand the pathophysiology of IVH and test pharmacological treatments. While there are existing models of neonatal IVH, those that reliably result in hydrocephalus are often limited by the necessity for large-volume injections, which may complicate modeling of the pathology or introduce variability in the clinical phenotype observed. Recent clinical studies have implicated hemoglobin and ferritin in causing ventricular enlargement after IVH. Here, we develop a straightforward animal model that mimics the clinical phenotype of PHH utilizing small-volume intraventricular injections of the blood breakdown product hemoglobin. In addition to reliably inducing ventricular enlargement and hydrocephalus, this model results in white matter injury, inflammation, and immune cell infiltration in periventricular and white matter regions. This paper describes this clinically relevant, simple method for modeling IVH-PHH in neonatal rats using intraventricular injection and presents methods for quantifying ventricle size post injection.

Effect of ornithokinin on feeding behavior, cloacal temperature, voluntary activity and crop emptying rate in chicks.

Bradykinin is a well-studied bioactive peptide associated with several physiological functions, including vasodilation and inflammation, in mammals. However, its avian homolog, ornithokinin, has received less research attention in birds. Therefore this study aimed to investigate the effect of intraperitoneal (IP) and intracerebroventricular (ICV) injections of ornithokinin on feeding behavior, cloacal temperature, voluntary activity, crop emptying rate, and blood constituents in chicks (Gallus gallus). We also investigated the effect of lipopolysaccharide (LPS), a cell wall component of gram-negative bacteria, on ornithokinin-associated gene expression was also investigated to determine whether activation of the ornithokinin system is induced by bacterial infection. Both IP and ICV injections of ornithokinin significantly decreased feed intake, cloacal temperature, voluntary activity, and crop emptying rate in chicks, but they did not affect the plasma concentration of corticosterone. Additionally, LPS significantly increased the expression of ornithokinin B2 receptor mRNA in several organs. Hence, ornithokinin is associated with a range of physiological responses in chicks and may be related to their response to bacterial infection.

Chronic intracerebroventricular infusion of angiotensin II causes dose- and sex-dependent effects on intake behaviors and energy homeostasis in C57BL/6J mice.

The renin-angiotensin system (RAS) within the brain is implicated in the control of fluid and electrolyte balance, autonomic functions, blood pressure, and energy expenditure. Mouse models are increasingly used to explore these mechanisms; however, sex and dose dependencies of effects elicited by chronic intracerebroventricular (ICV) angiotensin II (ANG II) infusion have not been carefully established in this species. To examine the interactions among sex, body mass, and ICV ANG II on ingestive behaviors and energy balance, young adult C57BL/6J mice of both sexes were studied in a multiplexed metabolic phenotyping system (Promethion) during chronic infusion of ANG II (0, 5, 20, or 50 ng/h). At these infusion rates, ANG II caused accelerating dose-dependent increases in drinking and total energy expenditure in male mice, but female mice exhibited a complex biphasic response with maximum responses at 5 ng/h. Body mass differences did not account for sex-dependent differences in drinking behavior or total energy expenditure. In contrast, resting metabolic rate was similarly increased by ICV ANG II in a dose-dependent manner in both sexes after correction for body mass. We conclude that chronic ICV ANG II stimulates water intake, resting, and total energy expenditure in male C57BL/6J mice following straightforward accelerating dose-dependent kinetics, but female C57BL/6J mice exhibit complex biphasic responses to ICV ANG II. Furthermore, control of resting metabolic rate by ANG II is dissociable from mechanisms controlling fluid intake and total energy expenditure. Future studies of the sex dependency of ANG II within the brain of mice must be designed to carefully consider the biphasic responses that occur in females.

Central administration of neuropeptide Y reduces the cellular heat stress response and may enhance spleen antioxidative functions in heat-exposed chicks.

Previously it was found that mRNA expression of neuropeptide Y (NPY) was increased in the chicken brain under heat stress. NPY has also been reported as an anti-stress factor to regulate brain functions in heat-exposed chicks. However, to the best of our knowledge, there is no report on the action of central NPY in the immune organs under heat stress. The aim of this study was to examine whether central injection of NPY can regulate heat stress response in the spleen and liver. After intracerebroventricular (ICV) injection of NPY, chicks were exposed to control thermoneutral temperature (CT: 30 ± 1 °C) or high ambient temperature (HT: 35 ± 1 °C) chambers for 60 min. Central injection of NPY caused lowering in rectal temperature under CT, but not under HT. Moreover, ICV injection of NPY caused a significant lower mRNA expression of heat-shock protein-70 and higher expression of glutathione synthase in the spleen, but not liver. Furthermore, plasma uric acid concentrations were significantly increased by the ICV injection of NPY in chicks under HT. These results indicate that brain NPY may contribute to attenuate the intracellular heat stress response and enhance antioxidative status in the immune organ, spleen in chicks.

Intraventricular Drug Delivery and Sampling for Pharmacokinetics and Pharmacodynamics Study.

Although the blood-brain barrier (BBB) protects the brain from foreign entities, it also prevents some therapeutics from crossing into the central nervous system (CNS) to ameliorate diseases or infections. Drugs are administered directly into the CNS in animals and humans to circumvent the BBB. The present protocol describes a unique way of treating brain infections through intraventricular delivery of antibiotics, i.e., polymyxins, the last-line antibiotics to treat multi-drug resistant Gram-negative bacteria. A straightforward stereotaxic surgery protocol was developed to implant a guide cannula reaching into the lateral ventricle in rats. After a recovery period of 24 h, rats can be injected consciously and repeatedly through a cannula that is fitted to the guide. Injections can be delivered manually as a bolus or infusion using a microinjection pump to obtain a slow and controlled flow rate. The intraventricular injection was successfully confirmed with Evans Blue dye. Cerebrospinal fluid (CSF) can be drained, and the brain and other organs can be collected. This approach is highly amenable for studies involving drug delivery to the CNS and subsequent assessment of pharmacokinetic and pharmacodynamic activity.

Effect of sodium nitroprusside on feeding behavior, voluntary activity, and cloacal temperature in chicks.

Nitric oxide (NO) is a well-known gaseous signaling molecule that is involved in a variety of physiological and pathological processes in vertebrates. The role of NO in physiological responses of birds has been investigated primarily using NOS inhibitors. Therefore, the effect of the absence of NO is well characterized. However, there is little knowledge on the effects of abundant NO in birds, which is the case in birds that have infections. Therefore, the purpose of the present study was to determine if intraperitoneal (IP) and intracerebroventricular (ICV) injections of sodium nitroprusside (SNP), a NO donor, affected feed intake, voluntary activity, cloacal temperature, crop emptying rate, and blood constituents in domesticated chicks (Gallus gallus) as model birds. We found that both IP and ICV injections of SNP significantly decreased feed intake while there was little effect on voluntary activity. Cloacal temperature was temporarily, but significantly, decreased by both types of injection of SNP. Additionally, both IP and ICV injections of SNP significantly decreased the crop emptying rate. The IP injection of SNP significantly increased the plasma concentrations of NO2/NO3, which are metabolites of NO, and corticosterone, and decreased the plasma glucose concentrations, while the ICV injection had no effect. The IP injection of SNP also showed the tendency to increase the nitrotyrosine level, to increase superoxide dismutase activity, and to decrease catalase activity in the plasma. These results suggest that under specific situations which produce abundant NO such as infection, NO would induce anorexia, hypothermia, inhibition of feed passage, and activation of the hypothalamus-pituitary-adrenal axis in chicks.

AMPA and angiotensin type 1 receptors are necessary for hemorrhage-induced vasopressin secretion.

Hypovolemia induced by hemorrhage is a common clinical complication, which stimulates vasopressin (AVP) secretion by the neurohypophysis in order to retain body water and maintain blood pressure. To evaluate the role of brain L-glutamate and angiotensin II on AVP secretion induced by hypovolemia we induced hemorrhage (∼25% of blood volume) after intracerebroventricular (icv) administration of AP5, NBQX, or losartan, which are NMDA, AMPA, and AT1 receptor antagonists, respectively. Hemorrhage significantly increased plasma AVP levels in all groups. The icv injection of AP5 did not change AVP secretion in response to hemorrhage. Conversely, icv administration of both NBQX and losartan significantly decreased plasma AVP levels after hemorrhage. Therefore, the blockade of AMPA and AT1 receptors impaired AVP secretion in response to hemorrhage, suggesting that L-glutamate and angiotensin II acted in these receptors to increase AVP secretion in response to hemorrhage-induced hypovolemia.

Delivery of Antisense Oligonucleotides to the Mouse Brain by Intracerebroventricular Injections.

The use of antisense oligonucleotides (AONs) is a promising therapeutic strategy for central nervous system disorders. However, the delivery of AONs to the central nervous system is challenging because their size does not allow them to diffuse over the blood-brain barrier (BBB) when injected systemically. The BBB can be bypassed by administering directly into the brain. Here we describe a method to perform single and repeated intracerebroventricular injections into the lateral ventricle of the mouse brain.

Management skin manifestation of multisystem inflammatory syndrome associated with SARS-CoV-2.

BACKGROUND: Multisystem inflammatory syndrome (MIS), which develops after a past covid-19 infection. MIS can be described in different tissue inflammation, including the heart, lung, kidney, brain, skin, eye, and or gastrointestinal organs at the presence of COVID-19. Initially, MIS was described in Europe in children infected with SARS-CoV-2, then it was recently seen in the USA in 2020. MIS is a rare but serious disease condition associated with COVID-19 that can affect children (MIS-C) and adults (MIS-A). CASE PRESENTATION: A 44-year-old male who showed MIS-A in 59-day after his first covid-19 contact history. The patient presented to our emergency department with complaints of high fever, nausea, weakness, redness of the eyes, headache, and joint pain. On the second day of his hospitalization, a maculopapular skin lesion was seen in most of the skin. His fever could not be controlled even given paracetamol and broad effective antibiotics. His clinical, radiological, and laboratory findings showed that he had MIS-A. The patient was given intravenous pulse methylprednisolone and intravenous immunoglobulin (IVIG). These treatments, then, resulted in improvement of his clinical conditions, including fever and skin lesions, on the second day of the treatment. The patient was discharged in 14 days after the treatment. CONCLUSION: This report indicated that diagnosis and treatment of MIS-A could result in reducing patient morbidity and mortality.

Human in silico trials for parametric computational fluid dynamics investigation of cerebrospinal fluid drug delivery: impact of injection location, injection protocol, and physiology.

BACKGROUND: Intrathecal drug delivery has a significant role in pain management and central nervous system (CNS) disease therapeutics. A fluid-physics based tool to assist clinicians in choosing specific drug doses to the spine or brain may help improve treatment schedules. METHODS: This study applied computational fluid dynamics (CFD) and in vitro model verification to assess intrathecal drug delivery in an anatomically idealized model of the human CSF system with key anatomic features of the CNS. Key parameters analyzed included the role of (a) injection location including lumbar puncture (LP), cisterna magna (CM) and intracerebroventricular (ICV), (b) LP injection rate, injection volume, and flush volume, (c) physiologic factors including cardiac-induced and deep respiration-induced CSF stroke volume increase. Simulations were conducted for 3-h post-injection and used to quantify spatial-temporal tracer concentration, regional area under the curve (AUC), time to maximum concentration (Tmax), and maximum concentration (Cmax), for each case. RESULTS: CM and ICV increased AUC to brain regions by ~ 2 logs compared to all other simulations. A 3X increase in bolus volume and addition of a 5 mL flush both increased intracranial AUC to the brain up to 2X compared to a baseline 5 mL LP injection. In contrast, a 5X increase in bolus rate (25 mL/min) did not improve tracer exposure to the brain. An increase in cardiac and respiratory CSF movement improved tracer spread to the brain, basal cistern, and cerebellum up to ~ 2 logs compared to the baseline LP injection. CONCLUSION: The computational modeling approach provides ability to conduct in silico trials representative of CSF injection protocols. Taken together, the findings indicate a strong potential for delivery protocols to be optimized to reach a target region(s) of the spine and/or brain with a needed therapeutic dose. Parametric modification of bolus rate/volume and flush volume was found to have impact on tracer distribution; albeit to a smaller degree than injection location, with CM and ICV injections resulting in greater therapeutic dose to brain regions compared to LP. CSF stroke volume and frequency both played an important role and may potentially have a greater impact than the modest changes in LP injection protocols analyzed such as bolus rate, volume, and flush.

The NKCC1 ion transporter modulates microglial phenotype and inflammatory response to brain injury in a cell-autonomous manner.

The NKCC1 ion transporter contributes to the pathophysiology of common neurological disorders, but its function in microglia, the main inflammatory cells of the brain, has remained unclear to date. Therefore, we generated a novel transgenic mouse line in which microglial NKCC1 was deleted. We show that microglial NKCC1 shapes both baseline and reactive microglia morphology, process recruitment to the site of injury, and adaptation to changes in cellular volume in a cell-autonomous manner via regulating membrane conductance. In addition, microglial NKCC1 deficiency results in NLRP3 inflammasome priming and increased production of interleukin-1ß (IL-1ß), rendering microglia prone to exaggerated inflammatory responses. In line with this, central (intracortical) administration of the NKCC1 blocker, bumetanide, potentiated intracortical lipopolysaccharide (LPS)-induced cytokine levels. In contrast, systemic bumetanide application decreased inflammation in the brain. Microglial NKCC1 KO animals exposed to experimental stroke showed significantly increased brain injury, inflammation, cerebral edema and worse neurological outcome. Thus, NKCC1 emerges as an important player in controlling microglial ion homeostasis and inflammatory responses through which microglia modulate brain injury. The contribution of microglia to central NKCC1 actions is likely to be relevant for common neurological disorders.

Neural labeling and manipulation by neonatal intraventricular viral injection in mice.

This step-by-step protocol provides a fast and easy technique to label and/or genetically manipulate neural cells, achieved by intraventricular injection of viral vectors into neonatal mice under ultrasound guidance. Successful injection of adeno-associated viral vectors (AAV) induces neural transduction as fast as 3 days post injection (dpi) in both the central and peripheral nervous systems. Virally driven expression persists until early adulthood. The same setup enables injection of other viral vectors as well as intramuscular injection. For complete details on the use and execution of this protocol, please refer to Wang et al. (2021) and Brill et al. (2016).