Name: Musekraniotomi til intracerebroventrikulær injektion med fri hånd
Uploaded: 2024-01-12T12:45:00
Description: Watch this Scientific Journal Video about Mouse Craniotomy for Free-Hand Intracerebroventricular Injection at JoVE.com

mouse craniotomy for free-hand intracerebroventricular injection

Udførelse af intracerebroventrikulær injektion i mus ved hjælp af en frihåndstilgang

The delivery of experimental agents, such as drugs1, viruses2, or cells3, to the brain is often necessary for neuroendocrine research. If the agent does not readily cross the blood-brain barrier or the experimental objective is to specifically test the central effects of the agent, it is important to have a reliable method for delivering injections into the brain. Moreover, injection into the intracerebroventricular (ICV) space provides the opportunity to distribute the agent widely in the brain and provides a large target area, thus increasing the likelihood of successful injection2. 
A common method for making ICV injections involves the placement of a permanent indwelling cannula. In this approach, a stereotaxic frame is necessary to position the commercially available or custom-made cannula, as the cannula is glued or cemented in place. Often, upon recovery, a supraphysiological dose of angiotensin II is administered through the cannula, and if drinking behavior is immediately observed, then the cannula is considered correctly placed4. This approach has many advantages, including the ability to perform long-term infusion and the ability to inject the same animal multiple times; additionally, if angiotensin II is employed, correct placement can be confirmed prior to the administration of experimental compounds. However, there are some limitations to placing a permanent cannula, including the requirement for expensive equipment (stereotaxic frame), the possibility of damage to the cannula after placement (e.g., mice can chew on the cannula of a cage mate), and the possibility of infections around the permanent cannula. Single ICV injections can be made with the use of a stereotaxic frame3, which, although effective, requires substantial exposure to anesthesia and, thus, may obscure some acute physiological and behavioral effects of the treatment. Additionally, the placement of mice in a stereotaxic frame requires substantial training to achieve stable placement and prevent the rupturing of the ear canals.
Here, an established method for making free-hand injections in mice is described. This method is based on previous reports5,6. The advantages of this technique are that it is simple, rapid, and does not require specialized equipment such as a stereotaxic frame. As described below, this procedure involves manipulating a glass syringe relative to landmarks on the mouse head to make the injections, which can be done rapidly and, thus, requires only a few minutes of gas anesthesia on the experimental day.

Forfatter Spotlight: Indsigt, innovationer og fremtidige veje inden for reproduktiv neurokontrol 

Frihånds intracerebroventrikulære injektioner hos mus

We are interested in the neural control of reproductive function, particularly how the brain regulates the release of hormones that control the gonads, both in healthy conditions and in response to stress. The main advantage of this technique is that it is relatively simple and rapid. Delivering drugs into the lateral ventricle is a great way to cast a wide net in testing a signaling pathway to support more detailed follow-up experiments.There's still much to be learned about the neuroendocrine control of reproduction, including the molecular mechanisms for luteinizing hormone pulse generation, mechanisms for modulating pulse frequency during stress, and generation of the preovulatory LH surge. In the future, we are particularly interested in integrating disparate findings into models.

Watch this Scientific Journal Video about Mouse Craniotomy for Free-Hand Intracerebroventricular Injection at JoVE.com

Musekraniotomi til intracerebroventrikulær injektion med fri hånd

Begynd med at forberede dig på en kraniotomi ved at placere en ren bænkpude på arbejdsfladen. Tape næsekeglen på en isofluranfordamper fast på arbejdsfladen. Placer derefter silastisk slange på enden af en 18-gauge nål, og sørg for, at en millimeter af nålespidsen stikker ud.Øv nålebevægelse ved at markere et ark papir med et udgangspunkt for bregma, yderligere 0,2 millimeter sideværts, og lav et endeligt mærke, en millimeter kaudal til det andet mærke. Øv dig i at flytte nålen fra startpunktet til det andet mærke og derefter det sidste mærke, og sørg for, at bevægelsen kan gentages uden guider. Barber derefter hovedet på det bedøvede dyr.Placer musen på arbejdsfladen, hold hovedet i næsekeglen for at opretholde anæstesi og påfør øjensmøremiddel på begge øjne. Skrub nu injektionsstedet med sterilt gasbind dyppet i iodopløsning. Tør derefter området af med en alkoholskrubbepude.Gentag denne proces tre gange. Hold hovedet fast med den ikke-dominerende hånd, og læg musen fladt på arbejdsfladen. Træk den forberedte 18-gauge nål hen over hovedhuden langs midterlinjen i en rostral, kaudale retning for at identificere bregma.Flyt nu nålen to millimeter sideværts og en millimeter kaudal til injektionsstedet. Hold nålen lodret, skub den fast gennem huden og knoglen, indtil slangen flugter med huden. Træk nålen tilbage, drej og tryk gennem huden og knoglen igen, gentag processen for at skabe et lille hul i knoglen.Brug den stumpe nål til at kontrollere, om der er et tilstrækkeligt hul i knoglen, stort nok til at lade nålen passere igennem. Fjern musen fra næsekeglen. Fortsæt med at rense blod fra injektionsstedet ved hjælp af sterilt gaze.Placer musen i et bur, på et burvarmer, indtil vågen.

Research

JoVE Journal

Neuroscience

Mouse Craniotomy for Free-Hand Intracerebroventricular Injection

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. ...