NG2-GLIA—NEURON CONNECTOME IN THE OLFACTORY BULB

NG2-glia engage in direct interactions with neurons, yet the spatial organization, directionality and cellular specificity of this connectivity in the olfactory bulb (OB) remain largely unexplored. Here, we provide an initial map of the bidirectional connectome between NG2-glia and neurons in the OB using complementary monosynaptic viral tracing strategies.
To dissociate neuron-to-glia and glia-to-neuron connectivity, we combined genetically targeted retrograde and anterograde viral tracers with defined directionality. High-resolution confocal imaging was coupled with automated cell segmentation (Cellpose), and quantitative spatial, morphological and layer-specific analyses using custom Python pipelines.
Retrograde tracing revealed that neurons providing synaptic input to NG2-glia exhibit a broader rostro-caudal distribution than neurons identified through anterograde tracing. These presynaptic neurons exhibited larger soma sizes and layer-dependent morphological differences and were predominantly inhibitory interneurons expressing parvalbumin, calretinin, and vasoactive intestinal peptide.
In contrast, anterograde tracing showed that NG2-glia preferentially target DCX-positive neurons, likely corresponding to immature, newly generated neurons migrating from the subventricular zone. These postsynaptic neurons were primarily located within the granular cell layer and displayed significantly smaller soma sizes, consistent with an immature or developing neuronal phenotype.
Together, these results reveal a marked asymmetry in NG2-glia connectivity, with distinct spatial distributions, morphological characteristics, and cellular identities depending on the direction of information flow. This work provides an initial framework for understanding the glia-neuron wiring diagram in the OB and highlights NG2-glia as active, directionally selective elements within OB microcircuits.
Supported by grant PID2022-136882NB-I00, funded by MICIU/AEI/ 10.13039/501100011033.