Vagal afferent innervation of the proximal gastrointestinal tract mucosa: chemoreceptor and mechanoreceptor architecture

Abstract

The vagus nerve supplies low-threshold chemo- and mechanosensitive afferents to the mucosa of the proximal gastrointestinal (GI) tract. The absence of a full characterization of the morphology and distributions of these projections has hampered comprehensive functional analyses. In the present experiment, dextran (10K) conjugated with tetramethylrhodamine and biotin was injected into the nodose ganglion and used to label the terminal arbors of individual vagal afferents of both rats and mice. Series of serial 100-μm thick sections of the initial segment of the duodenum as well as the pyloric antrum were collected and processed with diaminobenzidine for permanent tracer labeling. Examination of over 400 isolated afferent fibers, more than 200 from each species, indicated that three vagal afferent specializations, each distinct in morphology and in targets, innervate the mucosa of the proximal GI tract. One population of fibers, the villus afferents, supplies plates of varicose endings to the apical tips of intestinal villi, immediately subjacent to the epithelial wall. A second type of afferent, the crypt afferent, forms subepithelial rings of varicose processes encircling the intestinal glands or crypts, immediately below the crypt-villus junction. Statistical assessment of the isolated fibers indicated that the villus arbors and the crypt endings are independent, issued by different vagal afferents. A third vagal afferent specialization, the antral gland afferent, arborizes along the gastric antral glands and forms terminal concentrations immediately below the luminal epithelial wall. The terminal locations, morphological features, and regional distributions of these three specializations provide inferences about the sensitivities of the afferents.

Figure 1

A vagal villus afferent innervating a duodenal villus. A: A neurite of a villus afferent coursing through the submucosa and between intestinal crypts (lower right of panel) and then projecting to the apex of a villus (seen in transverse profile) to ramify in the apical half of the villus into a terminal plate (seen in side profile) at the basal side of the epithelial wall. B: A higher-power view of the apical terminal plate produced by the villus afferent seen in panel A. C: A higher-power view of terminal processes that the villus afferent in panel A distributes just deep to the basal pole of the epithelial wall of the villus. D: A higher-power view (area designated by arrow in panel A) of the trajectory of the villus afferent as it enters the tissue section (from left), courses near but without any tight apposition to the basal pole of crypt, and then reverses course and begins to ascend between two crypts (also labeled in panel A) on a path into the villus. c, crypt. Scale bars = 50 µm in A; 10 µm in B–D.

Figure 2

Vagal villus afferents illustrating details of typical trajectories followed by these afferents. A: A low-power montage of a fiber coursing through the smooth muscle wall (extreme lower right of image), dividing at the point, and then issuing a secondary neurite that breaks into three collaterals directed into a cone-shaped villus. Two of the collaterals can be seen ascending to roughly the midpoint of the villus, where they begin to ramify into a terminal arbor. This terminal arbor is somewhat truncated in the present case, because the apical tip of this wedge- or cone-shaped villus is located in adjacent section, and the arbor that can be seen is located in a “shoulder” of the villus. (The third collateral, at the far left, has been interrupted, but continued into the apex of the villus on an adjacent tissue section. The fourth collateral, which separates even as it enters the submucosa, at the lower far right, courses out of plane to innervate a second villus on a neighboring section.) B: The base of the villus illustrated in panel A. In this illustration the DIC interference has been attenuated in order to give a clearer view of the pattern of division and the continuity of the collaterals of the villus afferent in the submucosal and crypt region of the mucosa. C,D: A villus afferent in the apical tip of a villus illustrating how individual afferent neurites course, without branching or with minimal branching (see bottom of lower power panel D), into the apical pole of a villus where they then ramify extensively to end in plates of varicosities as well as numerous distributed terminals (higher-power panel C). c, crypt. Scale bars = 25 µm in A,B; 10 µm in C; 20 µm in D.

Figure 3

Villus afferent termination patterns at high power. A: At higher power, from a perspective looking at the villus wall formed by the basal pole of epithelial cells (the “honeycomb” or “cobblestone” or “waffle” pattern that can be seen faintly in the DIC image is the typical pattern formed by the basal epithelium) the terminal plates of villus afferents consist of webs of varicosities and terminals. In side profile (i.e., parallel to the epithelial wall), depending on the magnification and depth of field as well as the density of the terminal plate, these endings can appear as relatively simply profiles (such as the one illustrated in panel B) or more tightly packed mats of terminals (such as the examples in Figs. 1B, 2C). B: A terminal of a villus afferent seen, in side perspective, coursing up from the lower right to terminate along the epithelial wall of a villus. C.D: Illustration of one pattern of villus afferent termination with potential functional implications. Panel C is a luminal surface view of a villus tip scanned in the plane of the brush border. As the image indicates, the epithelial wall is scored with trenches that have been discussed in terms of movement and mechanical forces which play on the villus. These trenches can also be seen in side profile in panel B. As panel D, a side view (same perspective as in panel B) of the villus wall, illustrates vagal villus afferent processes in some cases terminate at the trenches, thus potentially exposing the afferent to mechanical forces. To eliminate ambiguities that might be introduced by an extended depth-of-field projection, this image in panel D is a single optical plane. Scale bars = 10 µm in A,D; 25 µm in B,C.

Figure 4

Vagal crypt afferents. A: A low-power transverse section of the duodenal wall in which a crypt afferent ends encircling the necks (region immediately below the crypt-villus transition) of three adjacent crypts. B: A higher-power view, with the same orientation as that in panel A, of a crypt afferent innervating a single crypt. The fiber coils around the more basal region of the crypt, making some apparent contacts at that level, and then continues to the more luminal neck region of the crypt where it terminates more profusely. C: A crypt afferent innervating four adjacent crypts (upper part of panel) while ignoring other neighboring crypts (bottom of panel). In this view, a plane of view tangent to the circular muscle wall or at right angles with the view in panel A, the image is taken through the “neck” or region immediately below the crypt. The labeled crypt afferent can be seen repeatedly encircling the target crypts. D: A higher-power view of a crypt afferent encircling a single crypt (same orientation as the crypts in panel C). Additional elements of the neurite can be seen running on crypt-villus transitional floor (e.g., lower right) and forming rings around additional crypts (upper right; partially cropped and out of field). SbMuc, submucosa; SM, smooth muscle wall. Scale bars = 50 µm in A; 20 µm in B,D; 25 µm in C.

Figure 5

Vagal antral gland afferents. A: A low-power view of part of the terminal arbor of an afferent innervating the gastric mucosa. One of two second-order collaterals of a vagal afferent fiber (entering from lower right corner of plate) distributes to form an arbor of higher order processes that travel along the glandular epithelial walls of the antral mucosa immediately adjacent to the antral lumen (far left of image). The collaterals tend to form varicose terminals and in some cases lamellar processes along the basal surfaces of the epithelial walls of the antral glands. On reaching the epithelium that constitutes the luminal surface, the collaterals often form aggregates of terminal varicosities and swellings. B: A higher-power image of two luminal aggregates of varicosities taken from the case illustrated in panel A (the area of the aggregates is designated with arrows in panel A). C: A lower-power image illustrating part of the arbor of terminals of a collateral of an antral afferent. In this case, the afferent collateral can be seen bifurcating in the lamina propria before coursing into the glandular mucosa to arborize. Scale bars = 50 µm in A; 20 µm in B; 25 µm in C.

Figure 6

Vagal antral afferent terminals in the gastric mucosa. A: An antral afferent neurite travels (from bottom of panel) to arborize into a plate of terminals immediately below the luminal epithelium of the antrum (in this case, the lumen is just immediately above the top of the image, and the gastric glands are running vertically to secrete into the lumen). B: An aggregate of terminals of an antral afferent located within the glandular wall. In this example, and in all panels of this figure, the glands are oriented in a vertical direction, and the afferent terminals are distributed among the necks of the glands, not at the gastric lumen. As illustrated by this terminal, many of the deeper terminal endings of the antral afferents–particularly those elements of the neurite that coursed laterally or at right angles to the length of the gastric glands–were commonly lamelliform or flattened in appearance. C: An example of the lamellar or growth-cone shaped terminals often formed by the antral afferents. Scale bars = 20 µm in A; 15 µm in B,C.