HD tissue specimens (n = 10) were collected at the time of pull-through surgery, while normal controls were obtained at the time of colostomy closure in patients with imperforate anus (n = 10). Nav1.9 immunofluorescence was visualized using confocal microscopy to assess the distribution of the protein. Western blot analysis was undertaken to determine Nav1.9 protein quantification.
Confocal microscopy revealed Nav1.9-immunoreactive neurons within the submucosal and myenteric plexus in normal controls, with a reduction in the HD specimens. Calbindin double-labeling showed that Nav1.9-immunoreactive neurons were IPANs. Nav1.9 channels were also seen to be co-localized on smooth muscle cells in all tissues. Western blotting revealed high levels of Nav1.9 protein expression in normal controls, while there was a marked decrease in Nav1.9 protein expression in the HD tissue.
Our results show the expression of Nav1.9 channels within the human colon for the first time. Furthermore, Nav1.9 channel expression is decreased in HD versus normal controls.