## 4 Discussion  
The present study was conducted to investigate the gut segment-specific effects of hyperglycemia and insulin treatment on myenteric 5-HT-IR neurons in STZ-induced type 1 diabetes. A single injection with a high dose of STZ leads to a rapid loss of pancreatic beta cells and concomitant hyperglycemia [37]. In this study, an average 25.5 mM blood glucose level was measured in diabetic rats with a less elevated body weight gain during the 10 weeks compared to controls. Moreover, STZ-induced diabetes is a widely used model of diabetic neuropathy with features including reduced size of nerve fibers, axon and myelin sheath, and decreased nerve conduction velocities [38]. The GI manifestation of neuropathy leads to GI motility disorders that were demonstrated in STZ-diabetic rodents including delayed gastric emptying of both liquids and solids, rapid small intestinal transit, and enhanced spontaneous activity of the colon [34; 39]. The proportion of 5-HT-IR hyperenteric neurons was about 2.5% in controls, which is in good agreement with previous results found in the guinea-pig ileum [17]. Moreover, we found no differences in the proportion of 5-HT-IR neurons between the investigated duodenum, ileum, and colon segments, which is in agreement with the immunohistochemical study of [PERSON] et al. [40] in which the localization, distribution, and projection of  
Figure 8: Total number of 5-HT-IR neurons in ganglia containing 5-HT-IR neurons. In controls, 1–2 serotonergic neurons were found in 5-HT-IR neurons containing myenteric ganglia. In diabetics, the most pronounced increase was found in the duodenum but the insulin treatment restored the level to close to that of the control. Data are expressed as mean \(\pm\) SEM. * \(p\) < 0.05 (between controls and diabetics); * \(p\) < 0.05 (between diabetics and insulin-treated diabetics). C—controls; D—diabetics; and ID—insulin-treated diabetics. 5-HT-IR neurons and fibers were similar in the stomach, duodenum, jejunum, ileum, and colon of rats. Although the role of 5-HT in GI motility under physiological conditions is controversial [18; 19], a growing amount of evidence demonstrates altered serotonergic signaling as an underlying mechanism in disorders with disturbed GI motility [25; 26] including diabetes mellitus [41; 42]. The effects of diabetes on the different neurochemical properties of myenteric neuronal populations have been widely investigated. The conclusion of those findings is an altered balance between inhibitory efferent (generally reduced) and excitatory (increased or unchanged) mechanisms [8; 39]. We demonstrated a significant 2.5-fold increase in the proportion of 5-HT-IR myenteric neurons in the duodenum and the colon and an increasing tendency in the ileum 10 weeks after STZ treatment application. Based on our earlier findings [33; 34] regarding changes in the total neuronal number in diabetes (unchanged in the duodenum and decreased in the ileum and colon), on present results of percentage of myenteric ganglia containing 5-HT-IR neurons, and the total number of serotonergic neurons in these ganglia, we presume that the increase in the percentage of 5-HT-IR neurons relative to the total neuronal number in the duodenum was due to neurochemical alterations of myenteric neurons. We found that more serotonergic neurons in 5-HT-IR ganglia and 5-HT-IR neurons were also detected in those ganglia that previously did not contain 5-HT-IR neurons. The above listed results of studies investigating 5-HT-IR nerves in different segments of the diabetic gut support our findings that the alterations are segment-specific. Increased 5-HT content was found in the duodenum and was unchanged in the rest of the small and large intestines 5 weeks after the allocan-induced hyperglycemia [30]. Lower brightness of fluorescence in 5-HT-IR nerves in the ileum and increased 5-HT level in the proximal colon were detected 8 weeks after STZ-induced diabetes [29].