Development of a platform for in vivo, two-photon targeted, multi-patch-clamping of interneurons.

This project has two main objectives: (1) to develop automated multi-neuron patching technology; (2) to apply it to test hypotheses concerning how local circuits incorporating interneurons operate during brain function and/or dysfunction. To observe such circuits functioning in vivo, a novel engineering approach is needed.
Patch clamp physiology is the gold standard technique for monitoring single neuron function. Paired patch clamp recordings allow unambiguous measurement of the connectivity between two neurons. Although in vivo concurrent multi-patching has been conducted in neurons (Jouhanneau et al., 2015; Poulet and Petersen, 2008; van Welie et al., 2016), the skill and labor required for multi-patching poses a challenge (Kodandaramaiah et al., 2012). This challenge could be tackled by automating the process. However, current automated processes are conducted in a "blind" way. This leads to a very reduced thoughput for specific types of cells, such as interneurons. This issue can be solved by using two-photon microscopy to target fluorescently labeled neurons.
This project will involve extending the platform developed in Annecchino et al. (2017) to multi-patch in vivo targeted cells (interneurons) for the first time. Initially, the platform will be developed for in vitro experiments, as they provide a more controlled environment. Once accomplished, in vivo testing will commence. Unique challenges to be tackled in this stage include cell movement as a result of tissue deformation from micropipette movement.
References:
Annecchino, L., Morris, A., Copeland, C., Agabi, O., Chadderton, P. and Schultz, S. (2017). Robotic Automation of In Vivo Two-Photon Targeted Whole-Cell Patch-Clamp Electrophysiology. Neuron, [online] 95(5), pp.1048-1055.e3. Available at: https://www.ncbi.nlm.nih.gov/pubmed/28858615 [Accessed 5 Nov. 2018].
Jouhanneau, J., Kremkow, J., Dorrn, A. and Poulet, J. (2015). In Vivo Monosynaptic Excitatory Transmission between Layer 2 Cortical Pyramidal Neurons. Cell Reports, 13(10), pp.2098-2106.
Kodandaramaiah, S., Flores, F., Holst, G., Singer, A., Han, X., Brown, E., Boyden, E. and Forest, C. (2018). Multi-neuron intracellular recording in vivo via interacting autopatching robots. eLife, [online] 7. Available at: https://cdn.elifesciences.org/articles/24656/elife-24656-v1.pdf [Accessed 5 Nov. 2018].
Kodandaramaiah, S., Franzesi, G., Chow, B., Boyden, E. and Forest, C. (2012). Automated whole-cell patch-clamp electrophysiology of neurons in vivo. Nature Methods, [online] 9(6), pp.585-587. Available at: https://www.nature.com/articles/nmeth.1993 [Accessed 5 Nov. 2018].
Lodato, S. and Arlotta, P. (2015). Generating Neuronal Diversity in the Mammalian Cerebral Cortex. Annual Review of Cell and Developmental
Poulet, J. and Petersen, C. (2008). Internal brain state regulates membrane potential synchrony in barrel cortex of behaving mice. Nature, 454(7206), pp.881-885.
van Welie, I., Roth, A., Ho, S., Komai, S. and Häusser, M. (2016). Conditional Spike Transmission Mediated by Electrical Coupling Ensures Millisecond Precision-Correlated Activity among Interneurons In Vivo. Neuron, 90(4), pp.810-823.