PINP: A New Method of Tagging Neuronal Populations for Identification during In Vivo Electrophysiological Recording

Abstract

Neural circuits are exquisitely organized, consisting of many different neuronal subpopulations. However, it is difficult to assess the functional roles of these subpopulations using conventional extracellular recording techniques because these techniques do not easily distinguish spikes from different neuronal populations. To overcome this limitation, we have developed PINP (Photostimulation-assisted Identification of Neuronal Populations), a method of tagging neuronal populations for identification during in vivo electrophysiological recording. The method is based on expressing the light-activated channel channelrhodopsin-2 (ChR2) to restricted neuronal subpopulations. ChR2-tagged neurons can be detected electrophysiologically in vivo since illumination of these neurons with a brief flash of blue light triggers a short latency reliable action potential. We demonstrate the feasibility of this technique by expressing ChR2 in distinct populations of cortical neurons using two different strategies. First, we labeled a subpopulation of cortical neurons—mainly fast-spiking interneurons—by using adeno-associated virus (AAV) to deliver ChR2 in a transgenic mouse line in which the expression of Cre recombinase was driven by the parvalbumin promoter. Second, we labeled subpopulations of excitatory neurons in the rat auditory cortex with ChR2 based on projection target by using herpes simplex virus 1 (HSV1), which is efficiently taken up by axons and transported retrogradely; we find that this latter population responds to acoustic stimulation differently from unlabeled neurons. Tagging neurons is a novel application of ChR2, used in this case to monitor activity instead of manipulating it. PINP can be readily extended to other populations of genetically identifiable neurons, and will provide a useful method for probing the functional role of different neuronal populations in vivo.

Internal States and Behavioral Decision-Making: Toward an Integration of Emotion and Cognition

“…5F,G), indicating that their activity is required during this consummatory phase of an aggressive encounter, confirming and extending earlier loss-of-function studies (Lin et al 2011;Sano et al 2013;Yang et al 2013). 9 Finally, preliminary experiments using optogenetic phototagging (Lima et al 2009) to identify Esr1 þ neurons in multi-electrode recordings indicate that these cells exhibit increased spiking rates during attack ( Fig. 5I -J; R Remedios and DJ Anderson, unpubl.).…”

Optogenetic Approaches to Target Specific Neural Circuits in Post-stroke Recovery

“…Given the approximate laser power for stimulation, the frequency, duration, and intervals of stimulations should be determined empirically depending on the brain region and cell type of interest. For example, single action potentials in parvalbumin (PV) inhibitory neurons of the primary somatosensory cortex are achieved by brief (<10 ms), high powered (>10 mW) laser pulses [128,129], whereas longer (1 s), low-power (0.1-3 mW) laser pulses are sufficient to drive spiking in striatal neurons [130]. Depending on the type of opsin used, the stimulation frequency and interval should match the intrinsic photocycle for high stimulation efficiency and low opsin desensitization [76,131].…”

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“…Cellular identity can be subsequently determined via intracellular 1 , juxtacellular 2 or loose-patch 3 iontophoresis of dye, but these recordings cannot be pre-targeted to specific neurons in regions with functionally heterogeneous cell types. Fluorescent proteins can be expressed in a cell-type specific manner permitting visuallyguided single-cell electrophysiology [4][5][6] . However, there are many model systems for which these genetic tools are not available.…”

“…Fluorescent proteins can be expressed in a cell-type specific manner permitting visuallyguided single-cell electrophysiology [4][5][6] . However, there are many model systems for which these genetic tools are not available.…”