Intrinsic sensory neurons provide direct input to motor neurons and interneurons in mouse distal colon via varicose baskets

J Comp Neurol. 2020 Aug;528(12):2033-2043. doi: 10.1002/cne.24872. Epub 2020 Feb 23.


Connections from intrinsic primary afferent neurons (IPANs), to ascending motor and interneurons have been described in guinea pig colon. These mono- and polysynaptic circuits may underlie polarized motor reflexes evoked by local gut stimulation. There is a need to translate findings in guinea pig to mouse, a species increasingly used in enteric neuroscience. Here, mouse distal colon was immunolabeled for CGRP, a marker of putative IPANs. This revealed a combination of large, intensely immunofluorescent axons in myenteric plexus and circular muscle, and thinner varicose axons with less immunofluorescence. The latter formed dense, basket-like varicosity clusters (CGRP+ baskets) that enveloped myenteric nerve cell bodies. Immunolabeling after 4-5 days in organ culture caused loss of large CGRP+ axons, but not varicose CGRP+ fibers and CGRP+ baskets. Baskets were characterized further by triple labeling with CGRP, nitric oxide synthase (NOS) and calretinin (CALR) antibodies. Approximately half (48%) of nerve cell bodies inside CGRP+ baskets lacked both NOS and CALR, while two overlapping populations containing NOS and/or CALR comprised the remainder. Quantitative analysis revealed CGRP+ varicosities were most abundant in baskets, followed by CALR+ varicosities, with a high degree of colocalization between the two markers. Few NOS+ varicosities occurred in baskets. Significantly higher proportions of CALR+ and CGRP+ varicosities colocalized in baskets than in circular muscle. In conclusion, CGRP+ baskets in mouse colon are formed by intrinsic enteric neurons with a neurochemical profile consistent with IPANs and have direct connections to both excitatory and inhibitory neurons.

Keywords: CGRP; RRID:AB_10000320; RRID:AB_2313775; RRID:AB_2314957; calretinin; colon; immunohistochemistry; motor neurons; mouse; nitric oxide synthase; primary afferent neurons.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Colon / innervation*
  • Enteric Nervous System / cytology*
  • Interneurons / cytology*
  • Mice
  • Mice, Inbred C57BL
  • Motor Neurons / cytology*
  • Neural Pathways / cytology
  • Sensory Receptor Cells / cytology*