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Despite numerous studies on the enteric nervous system (ENS), we lack fundamental knowledge on neuronal densities or total neuron numbers in different species. There are more anecdotal than actual figures on nerve counts.We used standardized preparation techniques and immunohistochemistry with validated panneuronal markers (human or mouse anti-HuD/C) to determine neuronal densities in specimen from the entire gastrointestinal tract of mice, guinea pig, and humans. In parallel, we measured the dimensions of the gastrointestinal regions in mouse and guinea pig. For humans, we had to rely on literature data.The average neuronal densities along the gastrointestinal tract were 35,011 ± 25,017 1/cmThis study reports the first comprehensive nerve cell count in mice, guinea pig, and human ENS. Neuronal densities were comparable between the three species and the differences in the total numbers of enteric neurons are likely due to body size and intestinal length. The number of enteric neurons is comparable to the number of neurons in the spinal cord for all three species.

P2X7 receptors play an important role in cytokine release and immune cell regulation. Their upregulation has been described in inflammatory and degenerative processes and P2X7 blockade or deletion has been shown to reduce tissue damage and severity of symptoms in animal models of inflammatory bowel disease (IBD). Several studies have found that P2X7 receptors are present on enteric neurons and glia and it was proposed that they mediate neuronal death during IBD. However, the cell type-specific localization of P2X7 receptors has been a matter of debate, since some antibodies have been found to be unspecific. Here we describe the preparation of whole-mount myenteric plexus from the colon of BAC transgenic P2X7-EGFP reporter mice and subsequent immunofluorescence staining of P2X7 receptors together with cell type-specific marker proteins.

Introduction: Investigating functions of neurons in the enteric nervous system using imaging techniques (e.g. voltage sensitive dyes) requires precise data concerning the three-dimensional structure of enteric ganglia. Recording from small enteric ganglia such as the submucosal plexus allows for single cell resolution recording, while myenteric ganglia are more complex and single cell resolution claim is not straightforward. Therefore, the aim of the present study was to analyze appositions of neuronal somata in myenteric ganglia. Material and methods: Stretched samples of guinea pig jejunum were fixed and dissected to expose the myenteric plexus. Tissues were stained for Hu protein and confocal microscopic analysis of myenteric ganglia was performed in order to reconstruct three-dimensional allocations of neuronal somata within the myenteric plexuses. Results: Myenteric neurons form a three-dimensional structure. Analysis of 103 cells showed that 20% were not overlapping with other cells, while 30% and 31% overlapped with 1 and 2 cells, respectively. Looking at the area of overlap, 42% of cells shared less than 10% of their area. On the other hand, 17% of cells had more than 50% of their area shared with others. Conclusions: These results show that in guinea pig myenteric ganglia neuronal somata form a monolayer, but up to half of the cells may have a significant overlap with other cell somata. Therefore all optical studies claiming single cell resolution recorded from the myenteric plexus should be confirmed by confocal analysis of the ganglia using immunohistochemical identification of cellular structure of analyzed plexuses.

Constipation and fecal impaction are frequent and distressing complaints in pediatric gastroenterology. Especially in neurologically handicapped children, treatment of severe forms of slow-transit constipation (STC) can be difficult. In the majority of cases, STC is of unknown etiology. However, in recent years, there is growing evidence that interstitial cells of Cajal (ICCs), which serve as electrical pacemakers and generate spontaneous electrical slow waves in the gastrointestinal tract, might play an important role in the pathophysiology of STC. It remains unclear whether morphological ICC alterations seen in affected patients are based on congenital developmental anomalies, or whether they are a consequence of long-term constipation with secondary damage of the gastrointestinal nervous system. To the best of our knowledge, we present the first case of a patient with histological alterations in ICC morphology who displayed multiple alterations of c-kit at the level of mRNA. The protein encoded by c-kit is the receptor tyrosine kinase Kit (CD117), which is crucial for development and function of ICCs. Therefore, these findings provide a new explanation for congenital alterations of ICC development that result in gastrointestinal motility disorders.