Injection of Adeno-Associated Virus Containing Optogenetic and Chemogenetic Probes into the Neonatal Mouse Brain
Optogenetics and chemogenetics are neuromodulation techniques used to study neuronal pathways. Viral vectors containing optogenetic or chemogenetic probes require at least a week after injection to produce effective expression levels in neurons. Therefore, injections need to be done in neonatal mice to enable experiments that require mice younger than 30 days of age. Here, we describe a protocol for performing surgery on neonatal mice and brain injection of a viral vector. The procedure utilizes cryoanesthesia and a pressure injector with a micropipette that can be directly injected through the skull of neonatal mice. Compared to other approaches, this protocol is relatively easy to implement and takes only minutes to perform, additionally allowing for increased numbers of injections. In the examples shown in this chapter, viral vectors were successfully delivered into the auditory cortex and the hippocampus as indicated by labeled expression of the optogenetic/chemogenetic probes. This technique provides a method for investigators to perform surgical injections and optogenetic/chemogenetic experiments in neonatal mice.
Key wordsOptogenetics Chemogenetics Surgery Rodent Neonatal mouse Cryoanesthesia Adeno-associated virus
The research was supported by research grant DC013073 from the National Institutes of Health.
- 4.Klapoetke NC, Murata Y, Kim SS, Pulver SR, Birdsey-Benson A, Cho YK, Moriomoto TK, Chuong AS, Carpenter EJ, Tian Z, Wang J, Xie Y, Yan Z, Zhang Y, Chow BY, Surek B, Melkonian M, Jayaraman V, Constantine-Paton M, Wong GK, Boyden ES (2014) Independent optical excitation of distinct neural populations. Nat Methods 11(3):338–346CrossRefGoogle Scholar
- 9.Armbruster B, Roth B (2005) Creation of designer biogenic amine receptors via directed molecular evolution. In: Neuropsychopharmacology, vol 30. Nature Publishing Group, London, pp S265–S265Google Scholar
- 11.Vardy E, Robinson JE, Li C, Olsen RH, DiBerto JF, Giguere PM, Sassano FM, Huang X, Zhu H, Urban DJ, White KL, Rittiner JE, Crowley NA, Pleil KE, Mazzone CM, Mosier PD, Song J, Kash TL, Malanga CJ, Krashes MJ, Roth BL (2015) A new DREADD facilitates the multiplexed chemogenetic interrogation of behavior. Neuron 86(4):936–946CrossRefGoogle Scholar
- 16.Chavkin C, Sud S, Jin W, Stewart J, Zjawiony JK, Siebert DJ, Toth BA, Hufeisen SJ, Roth BL (2004) Salvinorin A, an active component of the hallucinogenic sage Salvia divinorum is a highly efficacious κ-opioid receptor agonist: structural and functional considerations. J Pharmacol Exp Ther 308(3):1197–1203CrossRefGoogle Scholar
- 17.Jann MW, Lam YW, Chang WH (1994) Rapid formation of clozapine in guinea-pigs and man following clozapine-N-oxide administration. Archiv Int Pharmacodyn Ther 328(2):243–250Google Scholar
- 26.National Research Council (2003) Guidelines for the care and use of mammals in neuroscience and behavioral research. National Academies Press, Washington, DCGoogle Scholar
- 27.Kim JY, Grunke SD, Levites Y, Golde TE, Jankowsky JL (2013) Intracerebroventricular viral injection of the neonatal mouse brain for persistent and widespread neuronal transduction. J Vis Exp (91):51863–51863Google Scholar
- 33.Optogenetics Material Request/FAQ (n.d.) Optogenetics Resource Center. https://web.stanford.edu/group/dlab/optogenetics/request_dna.html