Expression Level of Hand2 Affects Specification of Enteric Neurons and Gastrointestinal Function in Mice

Abstract

Background & Aims Hand2 is a basic helix-loop-helix transcription factor required for terminal differentiation of enteric neurons. We studied Hand2 haploinsufficient mice, to determine whether reduced expression of Hand2 allows sufficient enteric neurogenesis for survival but not for development of a normal enteric nervous system (ENS). Methods Enteric transcripts that encode Hand2 and the neuron-specific embryonic lethal abnormal vision proteins HuB, HuC, and HuD were quantified. Immunocytochemistry was used to identify and quantify neurons. Apoptosis was analyzed with the TUNEL procedure. Intracellular microelectrodes were used to record inhibitory junction potentials. Gastrointestinal transit and colonic motility were measured in vivo. Results Levels of of enteric Hand2 transcripts were associated with genotypes of mice, in the following order: Hand2+/+ > Hand2LoxP/+ > Hand2+/− > Hand2LoxP/−. Parallel reductions were found in expression of HuD and, in regional and phenotypic manners, numbers of neurons; numbers of nNOS+ and calretinin+, but not substance P+ or vasoactive intestinal peptide+ neurons, decreased. No effects were observed in stomach or cecum. Apoptosis was not detected, consistent with the concept that Hand2 inhibits neuronal differentiation, rather than regulates survival. The amplitude of inhibitory junction potentials in colonic circular muscle was similar in Hand2 wild-type and haploinsufficient mice, although in haploinsufficient mice, the purinergic component was reduced and a nitrergic component appeared. The abnormal ENS of haploinsufficient mice slowed gastrointestinal motility but protected mice against colitis. Conclusion Reduced expression of factors required for development of the ENS can cause defects in the ENS that are subtle enough to escape detection yet cause significant abnormalities in bowel function.

Regulatory Logic Underlying Diversification of the Neural Crest

“…Ascl1 also has been shown to downregulate Sox10 , which is important for ensuring production of neurons [92]. Mutations in Hand2 cause downregulation of neuronal differentiation leading to a loss of of nNOS, calretinin, and VIP neuronal subtypes in mice [89,93,94]. Recently, an elegant paper has characterized the neuronal and glial profile of the mammalian gut [95].…”

Building a second brain in the bowel

“…Hand2 mutations reduce NCAM1, among many other defects. For example, HAND2 is needed for terminal differentiation of enteric neurons (12,87,88) and this more global effect on neurogenesis may underlie the effect of Hand2 mutations on ganglion formation and bowel motility. Additional work is needed to identify HAND2 targets and to define mechanisms controlling enteric ganglia formation.…”

Emerging complexity of the HuD/ELAVl4 gene; implications for neuronal development, function, and dysfunction

“…HuD transcripts are also present in several regions of the developing hippocampal formation, olfactory bulb, retina and spinal cord, such as sensory and motor neurons (Okano and Darnell 1997;Clayton et al 1998;). In the adult mouse and rodent nervous system, expression of HuD mRNA is mostly restricted to specific neuronal populations, including large pyramidal-like neurons in layer V of the neocortex and cerebellar cortex (Purkinje cells), the four Cornu Amonis (CA1-4) regions of the hippocampus, dorsal root ganglia, and motor neurons in the spinal cord, mitral cells in the olfactory bulb, ganglion and internal plexiform layers in the retina, and enteric nervous system neurons (Okano and Darnell 1997;Clayton et al 1998;D'Autreaux et al 2011).…”