The nervous system presents several unique challenges that make it a difficult system to study experimentally. At a structural level, the brain has a complexity that is orders of magnitude greater than other organs, and even the peripheral nervous system is profoundly complex. At a cellular level, neurons and accessory cells have extreme morphologies, physiological properties, and sensitivities that make them challenging to manipulate experimentally.

In addition to these difficulties, the brain is also protected by the so-called blood-brain-barrier (BBB). The endothelial cells forming the vessels of the brain are highly selective in regulating passage into the cerebrospinal fluid, preventing the entry of viruses and bacteria, while regulating the transport of hormones, ions, drugs, and other molecules. The BBB hampers the effectiveness of many in vivo experimental techniques. For example, most drugs and viral vectors delivered into the blood cannot effectively cross the BBB and infiltrate the brain.

The importance of CNS research has driven researchers to develop several approaches to circumvent the BBB and introduce viral vectors into the brain. The most obvious approach is direct injection of virus (AAV, lentivirus, etc.) into the extracellular fluid of the brain^{1, 2}. This method is highly effective in many circumstances, but is technically challenging, can be traumatic for the animal, and usually achieves poor distribution throughout the CNS. Intravenous injection of some types of viral vectors can be effective when combined with mannitol administration, which temporarily compromises the BBB³. Researchers have also identified specific virus serotypes that can cross the BBB adequately for many purposes. Most notably, AAV9 can cross the BBB by an active transport mechanism and effectively transduce multiple CNS cell types, including neurons and glia, although often requiring high viral load^{3, 4}.

Recently, a group of researchers at Caltech used a directed evolution approach they termed CREATE (Cre Recombinase-based AAV Targeted Evolution) to develop novel AAV capsid variants that can more efficiently deliver transgenes to the nervous system^5,6. Their new AAV serotypes are able to widely transduce adult mouse neurons and glia throughout the CNS and PNS (in the case of variant AAV-PHP.B), or specifically target cells of the CNS (with AAV-PHP.eB) or PNS (with AAV-PHP.S). These serotypes seem to be significantly more effective than AAV9, from which they are derived, achieving more than a 40-fold increase in gene transduction. For research and clinical applications where AAV9 is not effective or where high viral loads are problematic, these new AAV capsid variants may be great options for transduction of cells in the nervous system.

VectorBuilder

VectorBuilder offers a wide range of AAV capsid options, including the variants described here. Contact us to find out more, or with help designing your custom vectors. Use our award-winning online platform to design and order custom vectors specific to your research needs. Choose from AAVs, lentiviruses, adenovirus, shRNA expression vectors, CRISPR/Cas9 vectors, and more! Vectorbuilder also offers DNA preparation and virus packaging services, allowing you to focus on your experiments instead of making reagents.

References

1. Lowery RL, Majewska AK. Intracranial injection of adeno-associated viral vectors. J Vis Exp. 2010 Nov 17;(45). pii: 2140.
2. Kim JY, Grunke SD, Levites Y, Golde TE, Jankowsky JL. Intracerebroventricular viral injection of the neonatal mouse brain for persistent and widespread neuronal transduction. J Vis Exp. 2014 Sep 15;(91):51863.
3. Murlidharan G, Samulski RJ, Asokan A. Biology of adeno-associated viral vectors in the central nervous system. Front Mol Neurosci. 2014 Sep 19;7:76.
4. Gray SJ, Matagne V, Bachaboina L, Yadav S, Ojeda SR, Samulski RJ. Preclinical differences of intravascular AAV9 delivery to neurons and glia: a comparative study of adult mice and nonhuman primates. Mol Ther. 2011 Jun;19(6):1058-69.
5. Deverman BE, Pravdo PL, Simpson BP, Kumar SR, Chan KY, Banerjee A, Wu WL, Yang B, Huber N, Pasca SP, Gradinaru V. Cre-dependent selection yields AAV variants for widespread gene transfer to the adult brain. Nat Biotechnol. 2016 Feb;34(2):204-9.
6. Chan KY, Jang MJ, Yoo BB, Greenbaum A, Ravi N, Wu WL, Sánchez-Guardado L, Lois C, Mazmanian SK, Deverman BE, Gradinaru V. Engineered AAVs for efficient noninvasive gene delivery to the central and peripheral nervous systems. Nat Neurosci. 2017 Aug;20(8):1172-1179.