The enteric neural network in the proximal murine colon shows a regularly occurring hypoganglionic region, which is here characterized using anatomical and electrophysiological techniques. Staining with NADPH diaphorase, methylene blue and cuprolinic blue in standard whole mounts and three-dimensional gut preparations of the murine proximal colon consistently revealed two hypoganglionic areas that were surrounded by a dense clustering of enteric neurons. This irregularity in the ganglionic plexus was found to be present in mice of three different genetic backgrounds, as well as in rats. The lack of myenteric ganglia in these regions was associated with an absence of the longitudinal muscle layer, as shown in cross-sections. Histochemical identification of interstitial cells of Cajal in Kit^W-lacZ/+ transgenic mice showed Kit-positive cells oriented parallel to both muscle layers of the colon. Kit-positive cells oriented parallel to the longitudinal muscle layers were absent in the hypoganglionic area described. Electrical field stimulation (EFS) elicited TTX-sensitive inhibitory junction potentials (IJPs), which showed region-specific characteristics. The initial partly apamin-sensitive hyperpolarization was present in all parts of the murine colon, whereas a second sustained L-NNA-sensitive hyperpolarization was absent in the cecum and decreased from the proximal to the distal colon. Dissecting the hypoganglionic area from the surrounding tissue abolished the otherwise normal inhibitory neurotransmission to the circular muscle (1.6 ± 1.4 mV and 2.6 ± 1.7 mV for the fast and slow component of IJP amplitude in the hypoganglionic area vs. 16.5 [[plusm]] 1.9 mV and 23.7 ± 2.7 mV for the fast and slow component of IJP amplitude in the neuron-rich area respectively, P < 0.01, n = 6), while dissection of an area of identical size with an intact myenteric network showed normal inhibitory neurotransmission, indicating that the hypoganglionic area receives essential functional neural input from the neuron-rich surrounding tissue. In summary, in the murine and rat proximal colon, a constant and distinct hypoganglionic region is described, with important concomitant changes in local electrophysiology.