Fat deposition in the tunica muscularis and decrease of interstitial cells of Cajal and nNOS-positive neuronal cells in the aged rat colon

Abstract

Little is known about the time course of aging on interstitial cells of Cajal (ICC) of colon. The aim of this study was to investigate the change of morphology, ICC, and neuronal nitric oxide synthase (nNOS)-immunoreactive cells in the aged rat. The proximal colon of 344 Fischer rats at four different ages (6, 31, 74 wk, and 2 yr) were studied. The immunoreactivity of c-Kit, nNOS, anti-protein gene product 9.5, and synaptophysin were counted after immunohistochemistry. The c-kit, stem cell factor (ligand of Kit), and nNOS mRNA were measured by real-time PCR. c-Kit and nNOS protein were assessed by Western blot. Isovolumetric contractile force measurement and electrical field stimulation (EFS) were conducted. The area of intramuscular fat deposition significantly increased with age after 31 wk. c-Kit-immunoreactive ICC and nNOS-immunoreactive neurons and nerve fibers significantly declined with age. mRNA and protein expression of c-kit and nNOS decreased with aging. The functional study showed that the spontaneous contractility was decreased in aged rat, whereas EFS responses in the presence of atropine and L-NG-Nitroarginine methyl ester were increased in aged rat. In conclusion, the decrease of proportion of proper smooth muscle, the density of ICC and nNOSimmunoreactive neuronal fibers, and the number of nNOS-immunoreactive neurons during the aging process may explain the agingassociated colonic dysmotility.aging; colon; interstitial cells of Cajal; neuronal nitric oxide synthase; rats; inbred F344 AGING LEADS TO THE IMPAIRMENT of organ function as a result of accumulation of diverse deleterious changes throughout the cells and tissues (16). Constipation is a common problem in the elderly and known to be associated with multiple factors, such as decreased intestinal secretory epithelial function, abnormal colonic motility caused by enteric neurodegeneration, and reduced contractile response of smooth muscle cells with age (3-5).Enteric neurodegeneration involves the loss of enteric neuron, especially excitatory cholinergic neurons (2). In contrast, nitrergic myenteric neurons are known to be selectively spared in the aged (9, 36, 45). However, there was a contrary report that nitrergic neurons decreased in aged rats (43). Nitrergic enteric neurons, which release nitric oxide (NO) generated by catalysis of neuronal NO synthase (nNOS), act as nonadrenergic, noncholinergic inhibitory neuron and induce colonic smooth muscle relaxation (42). This NO is known to enhance transit of the rat colon by mediating descending relaxation, which in turn facilitates propulsion of the colonic contents (32). Nitrergic nerves of colon may be an important component of colonic motility.Interstitial cells of Cajal (ICC) also play important roles in gastrointestinal motility. ICC play as an electrical pacemaker and mediate both inhibitory and excitatory motor neurotransmissions, which manifest as electrical slow waves in smooth muscles. Consequently, ICC contribute to segmenting and peristaltic contractile activity (20,33). ICC have ...

Comparison of Changes in the Interstitial Cells of Cajal and Neuronal Nitric Oxide Synthase-positive Neuronal Cells With Aging Between the Ascending and Descending Colon of F344 Rats

“…Similar to what was found in a previous report, 14 the proportion of fat in 74-week old rats and 2-year old rats was significantly higher in the AC than in DC (P = 0.031 and P = 0.004, respectively) ( Fig. 2).…”

“…Since we have reported age-related changes in the AC of Fisher rats in a previous study, 14 the present study focused on investigating the differences of AC and DC. The results of our study demonstrate the following differences between the AC and DC: (1) fat deposition rarely occurred in the DC; (2) the expression of KIT in the submucosal plexus was higher in the DC than the AC and the expression of nNOS in the myenteric plexus was higher in the AC than the DC, and both decreased with age; and (3) the reduction of NO and its functional effect was more severe in the DC.…”

“…14 The tissue sections were treated with 3% hydrogen peroxide and nonspecific binding sites were blocked. Then, the sections were incubated with primary antibodies.…”

“…12,13 In addition, stem cell factors and KIT protein activity is closely related to the development and sustenance of ICCs, and their expression levels decrease with age in the rat ascending colon (AC). [14][15][16] Neuronal degeneration in the enteric nervous system is an important mechanism of age-related functional changes in the guts of older animals. 17 Dysfunction of intestinal muscles is associated with a reduction in enteric neuronal cells and an impaired response to direct stimuli in the myenteric plexus.…”

“…29 NO enhances colonic transit by mediating descending relaxation and facilitates propulsion of the contents of the colon. 30 Age-related loss of nNOS in rats was reported in the AC and mid-colon; 14,27 however, it has not been studied in the descending colon (DC) of the rat. Although the extent of neuronal loss differed in human and rats, the region-specific neuronal change in DC remains to be investigated.…”