Ultrastructural Characterization of The Output of VIP Expressing Interneurons in Mouse Barrel Cortex

by Xiaojuan Zhou
Doctoral thesis
Date of Examination:2017-05-15
Date of issue:2017-11-15
Advisor:Prof. Dr. Jochen F. Staiger
Referee:Prof.Dr. Carolin Wichmann
Referee:Prof. Dr. Thomas Dresbach
Referee:Prof. Dr. Siegrid Löwel
Referee:Prof. Dr. Tim Gollisch
Referee:Camin PhD Dean
Persistent Address: http://dx.doi.org/10.53846/goediss-6559

 

 

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Abstract

English

Based on this preliminary result, a quantitative immunoelectron microscopy study was adopted to comprehensively characterize the output of VIP interneurons across the whole laminae of mouse barrel cortex. Using VIPcre/YFP transgenic mice, by pre-embedding anti-YFP staining combined with DAB visualization, all VIP cells as well as their processes were stained in black. By systematically sampling the axonal boutons across all layers, it was found that their targets were preferentially dendrites (80%) but also spines (7%) and somata (13%). This distribution remained similar in layers II to V. In layer I, all of the targets were dendrites; in layer VI, innervation over the somata rose up to 39% although dendrites still represented the majority of the targets (58%). Post-embedding anti-GABA immunogold staining was used to find the GABAergic profiles among the targets. The staining efficacy in terms of sensitivity and specificity was evaluated using ROC analysis. Based on the ROC curves, the optimal cut-off points were determined by the method “closest point to (0,1)”. It generated the GABA-immunopositive thresholds for dendrites (spines) and somata at 11.1 and 8.5 gold grains/µm² respectively. Using the thresholds, the GABA-immunopositive rates among the subcellular targets were calculated: only 37% of the dendrites, 7% of the spines, and 26% of the somata showed above-threshold immunogold labeling. Among the targeted dendrites, around a third were GABA-immunopositive in layers I to IV, while in layers V and VI, the rates rose to 48% and 39% respectively. Statistical analysis showed a layer-dependent targeting existed: a higher proportion of innervation was localized on GABAergic dendrites in deep layers than in superficial layers. Besides the non-VIP positive neuronal targets, 14% of the axonal boutons formed self-innervating synapses on other VIP expressing subcellular structures, mainly on the dendrites. This self-innervation existed in all layers except layer I and had the highest incidence in layer IV. Occasionally, the boutons were found to contact the non-neuronal profiles like astrocytes and blood vessels. In sum, the study shows the main targets of VIP interneurons are dendrites with the majority belonging to non-GABAergic principal cells at subcellular level. The dendritic targeting is layer-dependent. The results disagree with the notion that VIP interneurons are interneuron-specific interneurons and argue against the uniform disinhibitory VIP-to-Martinotti cell circuit motif. Besides innervation on dendrites, VIP interneurons play an important role in regulating both pyramidal neurons and interneurons by targeting their somata in layer VI. In conclusion, the study provides the anatomical evidence for the diverse output of VIP interneurons and suggests multiple functionalities of VIP interneurons in cortical circuitry.
Keywords: Cortical circuit; Electron microscopy; GABA; Immunogold labeling; Vasoactive intestinal polypeptide
 
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