A combinatory genetic strategy for targeting neurogliaform neurons in the mouse basolateral amygdala.

The mouse basolateral amygdala (BLA) contains various GABAergic interneuron subpopulations, which have distinctive roles in the neuronal microcircuit controlling numerous behavioral functions. In mice, roughly 15% of the BLA GABAergic interneurons express neuropeptide Y (NPY), a reasonably characteristic marker for neurogliaform cells (NGFCs) in cortical-like brain structures. However, genetically labeled putative NPY-expressing interneurons in the BLA yield a mixture of interneuron subtypes besides NGFCs. Thus, selective molecular markers are lacking for genetically accessing NGFCs in the BLA. Here, we validated the NGFC-specific labeling with a molecular marker, neuron-derived neurotrophic factor (NDNF), in the mouse BLA, as such specificity has been demonstrated in the neocortex and hippocampus. We characterized genetically defined NDNF-expressing (NDNF+) GABAergic interneurons in the mouse BLA by combining the Ndnf-IRES2-dgCre-D transgenic mouse line with viral labeling, immunohistochemical staining, and electrophysiology. We found that BLA NDNF+ GABAergic cells mainly expressed NGFC neurochemical markers NPY and reelin (Reln) and exhibited small round soma and dense axonal arborization. Whole-cell patch clamp recordings indicated that most NDNF+ interneurons showed late spiking and moderate firing adaptation. Moreover, ∼81% of BLA NDNF+ cells generated retroaxonal action potential after current injections or optogenetic stimulations, frequently developing into persistent barrage firing. Optogenetic activation of the BLA NDNF+ cell population yielded both GABA- and GABA receptor-mediated currents onto BLA pyramidal neurons (PNs). We demonstrate a combinatory strategy combining the NDNF-cre mouse line with viral transfection to specifically target adult mouse BLA NGFCs and further explore their functional and behavioral roles.