Phenotype of Neurons in the Nucleus Tractus Solitarius Labeled using an Optogenetic Construct with a CaMKIIa Promoter




Lopez, Jackson D.


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Optogenetics is a very useful technique used to activate or inhibit specific cells of interest in specific brain regions. The purpose of this study was to determine the phenotype of cells within the nucleus tractus solitarius (NTS) of the rat using an optogenetic construct with a calcium calmodulin type II alpha type (CaMKIIa) promoter. Sixteen adult male rats were microinjected in the NTS with an optogenetic construct that consisted with an adeno-associated viral vector serotype 2 (AAV2) containing a CaMKIIa promoter, a light sensitive channelrhodopsin 2 (ChR2) and an mCherry reporter. Following a 3 to 4 week period after the microinjections, brain tissue was removed and sectioned. Immunohistochemistry was used to identify catecholaminergic neurons and astrocytes. In situ hybridization was used to identify neurons within the NTS that were glutamatergic or GABAergic. The vast majority of neurons that expressed the CaMKIIa promoter were catecholaminergic (87% in 4 rats), and 91% expressed the CaMKIIa enzyme. A subset of neurons expressing the CaMKIIa promoter were glutamatergic (38% in 5 rats). In contrast very few infected neurons were GABAergic (17% in 5 rats). The virus did not appear to infect astrocytes, because no GFAP immunoreactive cells expressed the CaMKIIa promoter. Based on these results, the optogenetic construct appears to be taken up by catecholaminergic neurons in the NTS with minimal or absent infection of GABAergic neurons or astrocytes. A subset of infected neurons were glutamatergic and future studies are needed to determine whether these neurons are also catecholaminergic.