S.M.H. was supported by postdoctoral fellowships from the Canadian Institutes for Health Research (CIHR), Heart &#38; Stroke Foundation of Canada (HSFC) and Alberta Innovates Health Solutions (AiHS); J.E.H. is supported by a grant from the National Heart, Lung and Blood Institute PO1HL-51971.

<ol>
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The authors have nothing to disclose.

Herein we have outlined, demonstrated and validated a novel method for targeted evaluation of RSNA in conscious mice, free to move and rest comfortably in their home cages. Following surgical implantation of an arterial pressure radiotelemeter, an indwelling intravenous infusion catheter and a custom-designed bipolar RSNA electrode, mice recovered from surgery and were left undisturbed for 48 to 72 hours. Mice remained comfortably settled in their home cage at all times (including experimental periods) with unrestricted ...

Interest in using mice in several areas of biomedical research continues to expand with the development of countless genetically engineered models. For the most part, technical advances have kept pace with the increased use of mice in physiology and there is now an impressive selection of miniaturized devices developed specifically for measuring important physiological parameters in mice. Although telemetric devices for direct measurement of autonomic nervous tone in the conscious rat have been available for over a decade, miniaturized devices for assessing nerve activity in conscious mice are currently not available. Investigators typically circumvent this limitation by evaluating the contribution of the autonomic nervous system with indirect methods (i.e. plasma or urine catecholamines, pharmacological autonomic blockade, spectral analysis of patterns of blood pressure/heart rate)1.
While these approaches provide valuable information, the result is a global picture of overall autonomic tone, rather than revealing the discrete contribution of isolated populations of nerves to the phenomenon under investigation. Alternatively, direct recording of activity from specific nerves has been executed in anesthetized mice, which poses a multitude of concerns. It is exceedingly difficult to maintain stable blood pressure within the physiological range in an anesthetized mouse for several hours following surgery. In fact, in these types of experiments, blood pressure is often unreported or presented at extremely low levels (i.e. 60-80 mmHg vs &#62;100mmHg in a conscious mouse)2. The fragility of the cardiovascular system exhibited in an anesthetized mouse preparation often precludes meaningful assessment of autonomic nerve activity, given the codependent relationship between blood pressure and sympathetic tone3,4.
To address this limitation, a new method for direct recording of renal sympathetic nerve activity (RSNA) in conscious, unrestrained mice, undisturbed within their home cages was developed. Both the surgical and experimental approach for successful implementation of this technique is described in detail. This preparation enables the investigator to simultaneously record arterial pressure via radiotelemetry in addition to RSNA, with the added capability to intravenously infuse agents of interest without disturbing the mouse.
Twenty four hours post-surgery, mice behave normally and do not exhibit signs of pain or distress. Experimental recordings may then commence 48 to 72 hours post-surgery while the mouse rests comfortably in its home cage with unrestricted access to food, water and environmental enrichment. Clear RSNA traces are presented and the characteristic responses of this nerve population to normal physical movements of the animal (such as eating and grooming) are demonstrated in addition to pharmacological modulation of systemic blood pressure. The quality and specificity of the RSNA signal is further validated by ganglionic blockade. This manuscript includes the audiovisual complement to an initially published description of this technique5.

<table><tbody><tr><td>Teflon-coated stainless steel multiple stranded wire</td><td>A-M Systems</td><td>793200</td><td>0.001in diameter bare; 0.0055in diameter coated</td></tr><tr><td>#11 Scalpel Blade</td><td>Fisher Scientific</td><td>ALMM9011</td><td></td></tr><tr><td>Soldering Iron and solder</td><td></td><td></td><td>Any make or model suitable</td></tr><tr><td>Male miniature pin connectors</td><td>A-M Systems</td><td>520200</td><td>Brass with gold plating</td></tr><tr><td>Female miniature pin connectors</td><td>A-M Systems</td><td>520100</td><td>Brass with gold plating</td></tr><tr><td>Heat Shrink tubing</td><td>Radio Shack</td><td>Model #: 278-1610 |&amp;nbsp;Catalog #: 2781610</td><td>1.6 mm diameter</td></tr><tr><td>Polyethylene 90 (PE90) tubing</td><td>VWR</td><td>CA-63018-703</td><td>0.86mm inner diameter; 1.27mm outer diameter</td></tr><tr><td>Dissecting microscope</td><td>Leica Microsystems</td><td>Leica M80</td><td>Any make or model also suitable</td></tr><tr><td>Polyethylene 10 (PE10) tubing</td><td>Braintree Scientific</td><td>PE10&amp;nbsp;50 FT</td><td>0.28mm inner diameter; 0.61mm outer diameter</td></tr><tr><td>Super Glue Liquid</td><td>Loctite</td><td>n/a</td><td>Liquid Formula; any brand suitable</td></tr><tr><td>Super Glue Gel</td><td>Loctite</td><td>n/a</td><td>Gel Formula; any brand suitable</td></tr><tr><td>Polyethylene tubing</td><td>Scientific Commodities</td><td>BB31695-PE/13</td><td>For pedestal 2.7mm inner diameter; 4.0mm outer diameter</td></tr><tr><td>Hospital Sterilization Services &amp;amp; Ozone Sterilization packets</td><td></td><td></td><td>Contact local hospital sterilization services</td></tr><tr><td>Isoflurane anesthesia</td><td>Abbott</td><td>05260-05</td><td></td></tr><tr><td>Deltaphase isothermal heat pads &amp;amp; surgical table</td><td>Braintree Scientific</td><td>39OP</td><td>Keep heat pads warm in a 37&amp;deg;C water bath; Corresponding surgical table essential</td></tr><tr><td>Glycopyrrolate</td><td>Amdipharm Mercury Company Limited</td><td>n/a</td><td></td></tr><tr><td>Isoflurane vaporizer system &amp;amp; flow gauge</td><td>Braintree Scientific</td><td>VP I</td><td>Include medical grade oxygen supply</td></tr><tr><td>Tissue scissors</td><td>Fine Science Tools</td><td>14173-12</td><td></td></tr><tr><td>Fine spring scissors</td><td>Fine Science Tools</td><td>15006-09</td><td></td></tr><tr><td>Small cotton-tipped applicators</td><td>Fisher Scientific</td><td>23400100</td><td></td></tr><tr><td>Fine Straight Forceps</td><td>Fine Science Tools</td><td>11254-20</td><td>#5, FST by Dumont Biologie Tip</td></tr><tr><td>Angled Forceps</td><td>Fine Science Tools</td><td>11251-35</td><td>#5/45 FST by Dumont</td></tr><tr><td>Small Absorbent Spears</td><td>Fine Science Tools</td><td>18105-03</td><td></td></tr><tr><td>Parafilm</td><td>Sigma Aldrich</td><td>BR701605&amp;nbsp;ALDRICH</td><td></td></tr><tr><td>Kwik-Sil 2 component Silicone Polymer</td><td>World Precision Instruments (WPI)</td><td>KWIK-SIL</td><td>Purchase extra specialized tips from WPI</td></tr><tr><td>5-0 Polysorb Suture</td><td>Tyco Healthcare</td><td>n/a</td><td></td></tr><tr><td>6-0 Silk Suture</td><td>Braintree Scientific</td><td>SUT-S&amp;nbsp;104</td><td>Deknatel brand, spool</td></tr><tr><td>Radiotelemetry Probe</td><td>Data Sciences International (DSI)</td><td>TA11-PAC10</td><td></td></tr><tr><td>Radiotelemetry Receiver</td><td>Data Sciences International (DSI)</td><td>PhysioTel RPC-1</td><td></td></tr><tr><td>Ambient Pressure Reference</td><td>Data Sciences International (DSI)</td><td>Apr-01</td><td></td></tr><tr><td>Pressure Output Adapter</td><td>Data Sciences International (DSI)</td><td>R11CPA</td><td></td></tr><tr><td>Rena Pulse Tubing</td><td>Braintree Scientific</td><td>RPT-040</td><td></td></tr><tr><td>Infusion Swivel</td><td>Instech Solomon</td><td>375/D/22</td><td></td></tr><tr><td>Swivel Support Arm &amp;amp; Mount</td><td>Instech Solomon</td><td>SMCLA</td><td></td></tr><tr><td>Polysulfone button&amp;nbsp;</td><td>Instech Solomon</td><td>LW62S/6</td><td></td></tr><tr><td>Stainless steel spring</td><td>Instech Solomon</td><td>PS62</td><td></td></tr><tr><td>Vetbond surgical adhesive</td><td>3M</td><td>n/a</td><td></td></tr><tr><td>Triple Antibiotic Ointment</td><td>Fougera</td><td>n/a</td><td></td></tr><tr><td>PowerLab 8 Channel Data Acquisition System &amp;amp; Software</td><td>ADInstruments</td><td>PowerLab 8/35</td><td></td></tr><tr><td>PVC Insulated Cable</td><td>Belden</td><td>PVC Audio Connection Cable 32 AWG</td><td></td></tr><tr><td>Preamplification Headstage</td><td>Dagan Corporation</td><td>Model 4002</td><td></td></tr><tr><td>Differential Amplifier</td><td>Dagan Corporation</td><td>EX4-400</td><td></td></tr><tr><td>Sodium Nitroprusside</td><td>Sigma Aldrich</td><td>71778-25G</td><td></td></tr><tr><td>Phenylephrine</td><td>Sigma Aldrich</td><td>P6126-5G</td><td></td></tr><tr><td>Sterile Physiological Saline 0.9% NaCl</td><td>Beckton Dickinson</td><td></td><td>Contact local hospital supplier</td></tr><tr><td>hexamethonium</td><td>Sigma Aldrich</td><td>H0879-5G</td><td></td></tr><tr><td>Stainless Steel top anti vibration table</td><td>n/a</td><td>n/a</td><td>Custom designed in-house; Solid steel plate on a benchtop is also suitable</td></tr><tr><td>Faraday cage</td><td>n/a</td><td>n/a</td><td>Custom designed and constructed in-house</td></tr><tr><td>Small animal hair trimmer</td><td>n/a</td><td>n/a</td><td>Drugstore, men&#039;s beard trimmer suitable</td></tr><tr><td>Dipilatory Cream</td><td>n/a</td><td>n/a</td><td>Veet brand, sensitive skin formula</td></tr><tr><td>10% Povidone Iodine</td><td>Purdue Products</td><td>Betadiene</td><td></td></tr><tr><td>70% Ethanol</td><td>n/a</td><td>n/a</td><td></td></tr><tr><td>Steel microretractors</td><td>n/a</td><td>n/a</td><td>Made in-house. Bend a steel paper clip &amp;amp; loop 4-0 silk to form a retractor</td></tr><tr><td>Hemostats</td><td>Fine Science Tools</td><td>13011-12</td><td></td></tr><tr><td>Heat Gun</td><td>Fisher Scientific</td><td>09-201-27</td><td></td></tr></tbody></table>

novel approach for simultaneous recording of renal sympathetic nerve activity and blood pressure with intravenous infusion in conscious, unrestrained mice.

Renal sympathetic nerves contribute significantly to both physiological and pathophysiological phenomena. Evaluating renal sympathetic nerve activity (RSNA) is of great interest in many areas of research such as chronic kidney disease, hypertension, heart failure, diabetes and obesity. Unequivocal assessment of the role of the sympathetic nervous system is thus imperative for proper interpretation of experimental results and understanding of disease processes. RSNA has been traditionally measured in anesthetized rodents, including mice. However, mice usually exhibit very low systemic blood pressure and hemodynamic instability for several hours during anesthesia and surgery. Meaningful interpretation of RSNA is confounded by this non-physiological state, given the intimate relationship between sympathetic nervous tone and cardiovascular status. To address this limitation of traditional approaches, we developed a new method for measuring RSNA in conscious, freely-moving mice. Mice were chronically instrumented with radio-telemeters for continuous monitoring of blood pressure as well as a jugular venous infusion catheter and custom-designed bipolar electrode for direct recording of RSNA. Following a 48-72 hour recovery period, survival rate was 100% and all mice behaved normally. At this time-point, RSNA was successfully recorded in 80% of mice, with viable signals acquired up to 4 and 5 days post-surgery in 70% and 50% of mice, respectively. Physiological blood pressures were recorded in all mice (116&plusmn;2 mmHg; n=10). Recorded RSNA increased with eating and grooming, as well-established in the literature. Furthermore, RSNA was validated by ganglionic blockade and modulation of blood pressure with pharmacological agents. Herein, an effective and manageable method for clear recording of RSNA in conscious, freely-moving mice is described.

Following the described protocol, survival rate was 100% - all mice instrumented in this study survived and recovered well following the surgical procedure. Within 24 hours of surgical preparation, all mice behaved normally, exhibiting typical eating, grooming and exploratory behaviors. No animals showed any sign of pain or distress at this time. 48 hours following surgery, a verifiable and clear RSNA signal was recorded in 10 out of the 12 mice. This signal was maintained in these mice 7...

Watch this Scientific Journal Video about Novel Approach for Simultaneous Recording of Renal Sympathetic Nerve Activity and Blood Pressure with Intravenous Infusion in Conscious, Unrestrained Mice. at

Novel Approach for Simultaneous Recording of Renal Sympathetic Nerve Activity and Blood Pressure with Intravenous Infusion in Conscious, Unrestrained Mice.

Anesthetized mice exhibit non-physiological systemic blood pressure, which precludes meaningful assessment of autonomic tone given the intimate relationship between blood pressure and the autonomic nervous system. Thus, a novel method to simultaneously record renal sympathetic nerve activity and blood pressure with intravenous infusion in conscious mice is outlined.

Renal sympathetic nerves contribute significantly to both physiological and pathophysiological phenomena. Evaluating renal sympathetic nerve activity ...

novel-approach-for-simultaneous-recording-renal-sympathetic-nerve

Research

JoVE Journal

Medicine

10.1K Views.  University of Alberta. Narcose muizen vertonen niet-fysiologische systemische bloeddruk, die zich verzet tegen de zinvolle beoordeling van autonome Toon gezien de intieme relatie tussen bloeddruk en het autonome zenuwstelsel. Dus, een nieuwe methode om tegelijkertijd record renale sympathiek zenuw activiteit en bloeddruk met intraveneuze infusie in de bewuste muizen wordt geschetst.

Video: Novel Approach for Simultaneous Recording of Renal Sympathetic Nerve Activity and Blood Pressure with Intravenous Infusion in Conscious, Unrestrained Mice.

Hyperinsulinemic-euglycemic Clamps in Conscious, Unrestrained Mice

Type 2 diabetes is characterized by a defect in insulin action. The hyperinsulinemic-euglycemic clamp, or insulin clamp, is widely considered the "gold standard" method for assessing insulin action in vivo. During an insulin clamp, hyperinsulinemia is achieved by a constant insulin infusion. Euglycemia is maintained via a concomitant glucose infusion at a variable rate. This variable glucose infusion rate (GIR) is determined by measuring blood glucose at brief intervals throughout the experiment and adjusting the GIR accordingly. The GIR is indicative of whole-body insulin action, as mice with enhanced insulin action require a greater GIR. The insulin clamp can incorporate administration of isotopic 2[14C]deoxyglucose to assess tissue-specific glucose uptake and [3-3H]glucose to assess the ability of insulin to suppress the rate of endogenous glucose appearance (endoRa), a marker of hepatic glucose production, and to stimulate the rate of whole-body glucose disappearance (Rd).
The miniaturization of the insulin clamp for use in genetic mouse models of metabolic disease has led to significant advances in diabetes research. Methods for performing insulin clamps vary between laboratories. It is important to note that the manner in which an insulin clamp is performed can significantly affect the results obtained. We have published a comprehensive assessment of different approaches to performing insulin clamps in conscious mice1 as well as an evaluation of the metabolic response of four commonly used inbred mouse strains using various clamp techniques2. Here we present a protocol for performing insulin clamps on conscious, unrestrained mice developed by the Vanderbilt Mouse Metabolic Phenotyping Center (MMPC; URL: www.mc.vanderbilt.edu/mmpc). This includes a description of the method for implanting catheters used during the insulin clamp. The protocol employed by the Vanderbilt MMPC utilizes a unique two-catheter system3. One catheter is inserted into the jugular vein for infusions. A second catheter is inserted into the carotid artery, which allows for blood sampling without the need to restrain or handle the mouse. This technique provides a significant advantage to the most common method for obtaining blood samples during insulin clamps which is to sample from the severed tip of the tail. Unlike this latter method, sampling from an arterial catheter is not stressful to the mouse1. We also describe methods for using isotopic tracer infusions to assess tissue-specific insulin action. We also provide guidelines for the appropriate presentation of results obtained from insulin clamps.

The hyperinsulinemic-euglycemic clamp, or insulin clamp, is the gold standard for assessing insulin action in vivo. A method for performing insulin clamps in mice is described. This includes a method for arterial catheterization that permits experiments to be performed in conscious, unrestrained mice with minimal stress.

Type 2 diabetes is characterized by a defect in insulin action. The hyperinsulinemic-euglycemic clamp, or insulin clamp, is widely considered the "gold ...

Femoral Arterial and Venous Catheterization for Blood Sampling, Drug Administration and Conscious Blood Pressure and Heart Rate Measurements

In multiple fields of study, access to the circulatory system in laboratory studies is necessary. Pharmacological studies in rats using chronically implanted catheters permit a researcher to effectively and humanely administer substances, perform repeated blood sampling and assists in conscious direct measurements of blood pressure and heart rate. Once the catheter is implanted long-term sampling is possible. Patency and catheter life depends on multiple factors including the lock solution used, flushing regimen and catheter material. This video will demonstrate the methodology of femoral artery and venous catheterization of the rat. In addition the video will demonstrate the use of the femoral venous and arterial catheters for blood sampling, drug administration and use of the arterial catheter in taking measurements of blood pressure and heart rate in a conscious freely-moving rat. A tether and harness attached to a swivel system will allow the animal to be housed and have samples taken by the researcher with minimal disruption to the animal. To maintain patency of the catheter, careful daily maintenance of the catheter is required using lock solution (100 U/ml heparinized saline), machine-ground blunt tip syringe needles and the use of syringe filters to minimize potential contamination. With careful aseptic surgical techniques, proper catheter materials and careful catheter maintenance techniques, it is possible to sustain patent catheters and healthy animals for long periods of time (several weeks).

Chronic catheterization of blood vessels in the rat is often required for administration of substances, obtain blood sample over a period of time or for direct conscious blood pressure measurements. Femoral arterial catheterization of the rat and corresponding measurements of blood pressure in the conscious animal will be demonstrated.

In multiple fields of study, access to the circulatory system in laboratory studies is necessary. Pharmacological studies in rats using chronically ...

Transcutaneous Assessment of Renal Function in Conscious Rodents

Glomerular filtration rate (GFR) is the gold standard to assess overall kidney function. However, traditional methods to evaluate GFR are cumbersome and time-consuming. In addition, serial blood or urine samples are required, with the associated stress for the experimental animals. A recent technique significantly reduces the investment in time and resources, minimizing the invasiveness and the animal stress, but being equally valid as the traditional approaches. The method measures transcutaneously renal function. Using an optical device and the exogenous renal marker fluorescein isothiocyanate (FITC)-sinistrin, this technique is capable of measuring the elimination kinetics of the marker through the skin. With neither blood nor urine samples nor the associated laboratory assays needed, the results of the transcutaneous measurement are almost instantaneously available. The method has been already validated in different species and successfully applied in several models of renal pathology. Moreover, due to its minimally invasive characteristics, it is suitable for sequential measurements within the same animal. Here is provided a detailed protocol to carry out the transcutaneous assessment of renal function in rodents.

Determination of glomerular filtration rate (GFR) is the gold standard to assess overall kidney function. However, traditional procedures to measure this parameter are cumbersome and require a large investment of time. Here we describe a faster and minimally invasive method to determinate GFR transcutaneously.

Glomerular filtration rate (GFR) is the gold standard to assess overall kidney function. However, traditional methods to evaluate GFR are cumbersome and ...

Quantifying Acute Changes in Renal Sympathetic Nerve Activity in Response to Central Nervous System Manipulations in Anesthetized Rats

Renal sympathetic nerve activity (RSNA) and mean arterial pressure are important parameters in cardiovascular and autonomic research; however, there are limited resources directing scientists in the techniques for measuring and analyzing these variables. This protocol describes the methods for measuring RSNA and mean arterial pressure in anesthetized rats. The protocol also includes the approaches for accessing the brain during RSNA recordings for central nervous system (CNS) manipulations. The RSNA recording technique is compatible with pharmacologic, optogenetic, or electrical stimulation of the CNS. The approach is useful when an investigator will measure short-term (min to h) autonomic responses in non-survival experiments to correlate anatomically with CNS nuclei. The approach is not intended to be used to obtain chronic (survival) recordings of RSNA in rats. Discharges in RSNA, averaged rectified RSNA, and mean arterial pressure can be quantified and analyzed further using parametric statistical tests. Methods for obtaining venous access, recording mean arterial pressure telemetrically, and brain fixation for future histological analysis are also described in the article.

Methods for measuring sympathetic and cardiovascular responses to central nervous system (CNS) manipulations are important for advancing neuroscience. This protocol was developed to assist scientists with measuring and quantifying acute changes in renal sympathetic nerve activity (RSNA) in anesthetized rats (non-survival).

Renal sympathetic nerve activity (RSNA) and mean arterial pressure are important parameters in cardiovascular and autonomic research; however, there are ...

Neuroscience

Simultaneous Electrocardiography Recording and Invasive Blood Pressure Measurement in Rats

For studies related to cardiovascular physiology or pathophysiology, blood pressure (BP) and electrocardiography are basic observational parameters. Research focusing on cardiovascular disease models, potential cardiovascular therapeutic targets or pharmaceutical agents requires assessment of systemic arterial pressure and heart rhythm changes. In situations where radio telemetry systems are not available or affordable, the technique of femoral artery cannulation is an alternative way to obtain intra-arterial pressure waveform recordings and systemic BP measurements. This technique is economical and can be performed with standard equipment in animal facilities. However, invasive arterial pressure recording requires cannulation of small arteries, which can be a challenging surgical skill. Here, we present step-by-step protocols for femoral artery cannulation procedures. Key procedures include the calibration of the data acquisition system, tissue dissection and femoral artery cannulation, and setup of the arterial cannulation system for pressure recording. Surface electrocardiography recording procedures are also included. We also present examples of BP recordings from normotensive and hypertensive rats. This protocol allows reliable direct recordings of systemic BP with simultaneous electrocardiography.

Here, we describe a setup for simultaneous recording of electrocardiography and intra-arterial blood pressure (BP) in experimental rats, which can be done with standard equipment in animal facilities and can be applied to physiological or pharmacological studies to investigate pathogenic or therapeutic mechanisms in cardiovascular medicine.

For studies related to cardiovascular physiology or pathophysiology, blood pressure (BP) and electrocardiography are basic observational parameters. ...

Implantation of Combined Telemetric ECG and Blood Pressure Transmitters to Determine Spontaneous Baroreflex Sensitivity in Conscious Mice

Blood pressure (BP) and heart rate (HR) are both controlled by the autonomic nervous system (ANS) and are closely intertwined due to reflex mechanisms. The baroreflex is a key homeostatic mechanism to counteract acute, short-term changes in arterial BP and to maintain BP in a relatively narrow physiological range. BP is sensed by baroreceptors located in the aortic arch and carotid sinus. When BP changes, signals are transmitted to the central nervous system and are then communicated to the parasympathetic and sympathetic branches of the autonomic nervous system to adjust HR. A rise in BP causes a reflex decrease in HR, a drop in BP causes a reflex increase in HR.
Baroreflex sensitivity (BRS) is the quantitative relationship between changes in arterial BP and corresponding changes in HR. Cardiovascular diseases are often associated with impaired baroreflex function. In various studies reduced BRS has been reported in e.g., heart failure, myocardial infarction, or coronary artery disease. 
Determination of BRS requires information from both BP and HR, which can be recorded simultaneously using telemetric devices. The surgical procedure is described beginning with the insertion of the pressure sensor into the left carotid artery and positioning of its tip in the aortic arch to monitor arterial pressure followed by the subcutaneous placement of the transmitter and ECG electrodes. We also describe postoperative intensive care and analgesic management. After a two-week period of post-surgery recovery long-term ECG and BP recordings are performed in conscious and unrestrained mice. Finally, we include examples of high-quality recordings and the analysis of spontaneous baroreceptor sensitivity using the sequence method.

The baroreflex is a heart-rate regulation mechanism by the autonomic nervous system in response to blood-pressure changes. We describe a surgical technique to implant telemetry transmitters for continuous and simultaneous measurement of electrocardiogram and blood pressure in mice. This can determine spontaneous baroreflex sensitivity, an important prognostic marker for cardiovascular disease.

Blood pressure (BP) and heart rate (HR) are both controlled by the autonomic nervous system (ANS) and are closely intertwined due to reflex mechanisms. ...

Long-Term Continuous Measurement of Renal Blood Flow in Conscious Rats

The kidneys play a crucial role in maintaining the homeostasis of body fluids. The regulation of renal blood flow (RBF) is essential to the vital functions of filtration and metabolism in kidney function. Many acute studies have been carried out in anesthetized animals to measure RBF under various conditions to determine mechanisms responsible for the regulation of kidney perfusion. However, for technical reasons, it has not been possible to measure RBF continuously (24 h/day) in unrestrained unanesthetized rats over prolonged periods. These methods allow the continuous determination of RBF over many weeks while also simultaneously recording blood pressure (BP) with implanted catheters (fluid-filled or by telemetry). RBF monitoring is carried out with rats placed in a circular servo-controlled rat cage that enables the unrestrained movement of the rat throughout the study. At the same time, the tangling of cables from the flow probe and arterial catheters is prevented. Rats are first instrumented with an ultrasonic flow probe placement on the left renal artery and an arterial catheter implanted in the right femoral artery. These are routed subcutaneously to the nape of the neck, and connected to the flowmeter and pressure transducer, respectively, to measure RBF and BP. Following surgical implantation, rats are immediately placed in the cage to recover for at least one week and stabilize the ultrasonic probe recordings. Urine collection is also feasible in this system. The surgical and post-surgical procedures for continuous monitoring are demonstrated in this protocol.

The present protocol describes a long-term continuous measurement of renal blood flow in conscious rats and simultaneously recording blood pressure with implanted catheters (fluid-filled or by telemetry).

The kidneys play a crucial role in maintaining the homeostasis of body fluids. The regulation of renal blood flow (RBF) is essential to the vital ...

Angled Silicon Multielectrode Recordings for Deep Brainstem Structures in Mice

Acute Single-Unit Multi-Electrode Recordings from the Brainstem of Head-Fixed Mice

Silicon multielectrode-based recordings are increasingly popular for studying neuronal activity at the temporal resolution of action potentials in many brain regions. However, recording neuronal activity from deep caudal structures like the brainstem using multi-channel probes remains challenging. A significant concern is finding a trajectory for probe insertion that avoids large blood vessels, such as the superior sagittal venous sinus and the transverse venous sinus. Injuring these large veins can cause extensive bleeding, damage to the underlying brain tissue, and potentially death. This approach describes targeting brainstem structures by coupling anterior coordinates with an angled approach, allowing the recording probe to penetrate the brain below high-risk vascular structures. Compared to a strictly vertical approach, the angled approach maximizes the number of brain regions that can be targeted. Using this strategy, the ventrolateral periaqueductal gray (vlPAG), a brainstem region associated with REM sleep, can be reproducibly and reliably accessed to obtain single-unit, multi-electrode recordings in head-fixed mice before and during sevoflurane anesthesia. The ability to record neuronal activity in the vlPAG and surrounding nuclei with high temporal resolution is a step forward in advancing the understanding of the relationship between REM sleep and anesthesia.

Author Spotlight: Investigating Anesthesia-Induced Sleep Pathways and Neuronal Excitability in Mice

This protocol describes a method to obtain in vivo, high-density single-neuron recordings from the brainstem of head-fixed mice. This approach is deployed to measure the action potential firing of neurons in the ventrolateral periaqueductal gray - a brainstem region inactive during Rapid Eye Movement (REM) sleep - before and during general anesthesia.

Silicon multielectrode-based recordings are increasingly popular for studying neuronal activity at the temporal resolution of action potentials in many ...

Recording Changes in Renal Sympathetic Nerve Activity in Response to Arterial Pressure in Rats

Source: Fink, A. M., et al. <a href="https://app.jove.com/v/58205">Quantifying Acute Changes in Renal Sympathetic Nerve Activity in Response to Central Nervous System Manipulations in Anesthetized Rats.</a> J. Vis. Exp. (2018).
This video demonstrates a technique of studying the activity of renal sympathetic nerves (RSNs) in response to changes in arterial pressure in rats. Upon attaching a pressure-sensing catheter to the femoral artery, the renal sympathetic nerves are exposed. The nerves are then lifted onto a recording electrode, and their electrical activity is recorded. Next, a vasoconstricting drug is injected to elevate blood pressure, and corresponding changes in RSN activity are monitored.

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. Create ...

JoVE Encyclopedia of Experiments

Neurophysiology

Electrophysiology Techniques

Sympathetic Denervation of Renal Arteries in an Angiotensin II-Induced Hypertension Mouse Model

Place the mouse on the operating table, keep the abdomen up, and fix the limbs with tape. Disinfect the abdominal skin with povidone iodine followed by 3 wipes with 70% ethanol. After making a 2 centimeter ventral midline abdominal incision, pull back the intestine with gauze soaked in 37 degrees Celsius saline.
To expose the left renal artery, carefully, but bluntly, dissect the fat away from the renal artery using curved tweezers. Using sterile sharp scissors, cut the weighing paper into a rectangle of the same size as the renal artery. Cut several pieces of the weighing paper at a time to keep the shape same.
Dip the weighing paper in 10% phenol ethanol solution for at least 30 seconds. Cover the surface of the left renal artery, and wrap the vessel with the weighing paper for 2 minutes. Use gauze to protect the surrounding tissues to avoid the weighing paper from touching the kidney and intestine.
Repeat the same procedure for the right renal artery. After surgery, reposition the muscles into their initial position and close the peritoneum with 6-0 vicryl sutures in an interrupted suture pattern. Then close the skin with interrupted 4-0 nylon sutures. Monitor all mice until fully recovered.

Novel Approach for Simultaneous Recording of Renal Sympathetic Nerve Activity and Blood Pressure with Intravenous Infusion in Conscious, Unrestrained Mice.

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