Spatial Distribution of R a t Fungif orm Papillae

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INGLIS J. MILLER, JR. AND ARTHUR J. PRESLAR Department of Anatomy, Bowman Gray School of Medicine of W a k e Forest University, Winston-Salem, North Carolina 271 03

ABSTRACT The objective of this study was to determine the spatial distribution of fungiform papillae on the rat's tongue. Since each fungiform papilla in the rat has a single taste bud, the spatial distribution of fungiform papillae is equivalent to the location of taste buds on the anterior tongue. A mean total number of 187 fungiform papillae per tongue were found which were about equally divided between the two lateral halves of the tongue. Over 50% of the total number of fungiform papillae were located on the tongue tip for an average density of 3.4 papillae/mm2, while the dorsal surface of the tongue had an average density of 1.3 papillae/mm2 of tongue surface. Papillae were absent on the dorsal midline, but a paracentral line of papillae running from anterior to posterior was a consistent finding. Though not identical, the distribution of papillae was essentially the same on different tongues. The functional significance of the papilla distribution is not understood, but electrophysiological experiments show evidence of neural interaction of papillae which are clustered together. The distribution of papillae and the distribution of nerve fibers which innervate them must be evaluated together in order to appreciate the significance of the distribution of fungiform papillae and their associated taste buds.

The gustatory papillae on the tongue of the Norway rat were identified and counted in a classic study by Fish et al. ('44) using an intravascular injection of methylene blue dye. With concurrent histological examination they showed that the fungiform papillae on the anterior two-thirds of the tongue contained a single taste bud. In the last two decades the rat fungiform papilla has been used extensively as a model system of the mammalian taste receptor in morphological and electrophysiological studies (Beidler, '71 ). Single chorda tympani neurons in the rat may receive input from more than one fungiform papilla (Miller, '71; Pfaffmann, '70) and simultaneous inputs to several fungiform papillae can evoke lateral depression or enhancement (Miller, '71, ' 7 2 ) . The fibers which innervate taste buds in the fungiform papillae branch and the branches interconnect adjacent papillae (Miller, '74). It is via these branches that the lateral interactions are believed to be mediated. Since the papillae that produce lateral interactions are clustered together on the ANAT. REC., 181: 679-684.

tongue surface, the object of this study was to quantify the similarity of papilla distributions in a population of rats and to identify regions of high papilla density where lateral interactions might be maximized by the contiguity of receptors. METHODS

Each of ten adult male Sprague-Dawley rats (275-325 gm) was anesthetized and secured ventral side up on a small animal operating board. The method of Fish et al. ('44) was used which employs the injection of about 30 ml of a 5% formalin in 0.1% methylene blue solution into the left ventricle of the heart during a period of about five minutes. The tongues were removed and photographed with a Leica M-2 camera back fitted with a Leitz adapter to a 5 x microscope eyepiece which was inserted into a Bausch and Lomb StereoReceived JuIy 15, '74. Accepted Nov. 7, '74. lThis work was supported in part by NIH Grant

NS 10389.

ZA preliminary report of these findings was presented at the Third Annual Meeting of the Society for Neuroscience, San Diego, CA, November, 1973.

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zoom 7 dissecting microscope. Photographs were made of all aspects of the tongue and printed on standard enlarging paper. The fresh tongues were measured to insure proper magnification factors and were preserved to allow visual examination to supplement the photographs in areas of low contrast, Photographs of the various aspects of the tongues were taped together to form a montage which had a h a 1 magnification of ten times the tongue’s width at its narrowest point. Each fungiform papilla was identified with a set of two coordinate numbers. A Cartesian coordinate system was applied to the montage so that the ordinate corresponded to the median sulcus of the tongue and the origin was located at the point of transition from the smooth epithelium of the ventral surface to the rough surface containing filiform papillae. The coordinates represented distances of 0.1 mm on the tongues. Ordinate numbers corresponded to anterior-posterior distances while abscissae corresponded to distances in the medial-buccal axis. RESULTS

Perfusion of the rat tongue with methylene blue solution produced a specimen as shown in figure 1. Fungiform papillae appeared as dark spots against a coarse gray background of filiform papillae. Panel a shows the dorsal surface; panel b shows the superior aspect of the tongue tip; and panel c shows the ventral aspect of the tongue tip. The dorsal midline is demarcated by a median dorsal sulcus which forms a prominent notch on the tip. To the left in panel a is a slightly elevated region called the intermolar eminence with coarse filiform papillae and no fungiform papillae. There is a transition in the surface between the smooth epithelium of the major portion of the ventral tongue surface and the rough texture of the surface containing filiform and fungiform papillae (fig. lc). A total of 1,871 fungiform papillae were found resulting in a mean ( * S.D.) of 187.1 C 9.2 papillae per tongue with a range of 175 to 201 among ten animals. An equal distribution of fungiform papillae was found on the two lateral halves of the

tongue with 93.0 C 5.5 papillae on the right sides and 95.9 -1- 7.6 papillae on the left sides. Only one tongue from among ten had an equal number of papillae on each side, while the greatest disparity among sides was 20 papillae. The locations of fungiform papillae are plotted for three tongues in figure 2 in which medial-lateral as well as anteriorposterior distances are shown in millimeters. The two halves of the tongue are generally bilaterally symmetrical although one side of the tongue is not the mirror image of the other half. Fungiform papillae are distributed over a surface 20 mm in length by about 10 mm in width, or about 200 mm2 of surface area. The highest density of papillae is located on the tip, while there is an absence of fungiform papillae along the midline of the middorsal region. Papillae on the medial margin of each half of the tongue form a conspicuous line of papillae from the anterior tip of the dorsal surface to a posterior limit at the intermolar eminence. These paracentral lines of papillae converge at the midline in the superior aspect of the tongue tip (fig. l c ) . The point of convergence of the paracentral lines of papillae corresponds approximately to the boundry between the dorsal and anterior aspects of the tongue tip. The distribution of papillae at various distances along the anterior-posterior axis is plotted in figure 3. There is an obvious mode at the tip of the tongues with means of 28.8 & 4.0 and 26.9 iz 4.9 papillae at distances 2 and 3 mm posterior of the origin, respectively, which grades off to a plateau from about 8 mm to about 17 mm posterior of the origin. The diminution of papillae from 18 to 20 mm results from slight differences in the lengths of the tongues. The filled circles in figure 3 indicate a cumulative percent of the total fungiform papillae along the anterior-posterior axis of the tongue. The anterior 4 mm of tongue surface contains 10.7%, 15.4% 14.4%, and 10.6%, respectively, of the total fungiform papillae of the tongue; so that 51.5% are located on the first 4 mm of the anterior and ventral aspects of the tongue tip. The average density of papillae

DISTRIBUTION OF RAT FUNGIFORM PAPILLAE

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Fig. 1 Photomicrographs of three aspects of a rat tongue perfused with methylene blue solution. Panel a shows the dorsal aspects of the entire tongue; panel b shows the dorsal aspect of the tongue tip; and panel c shows the ventral aspect of the tongue tip. Fungiform papillae appear as dark spots while fdiform papillae appear gray. The intermolar eminence shows at the left of panel a. A filled circle in panel c identifies the location of the origin of the coordinate system used to designate papillae. Approximate magnscation: x 5.4.

on the tongue tip is 3.4 papillae/mm', while the average density on the dorsal surface is 1.3 papillae/mm2. The distribution of fungiform papillae along the medial-lateral axis of the tongue is illustrated in figure 4. The graph for the entire tongue shows a conspicuous mode between 0.6 and 1.0 mm lateral to the mid-

line which corresponds to the paracentral line of papillae shown in figures 1 and 2. This mode is most conspicuous in the midregion but is also discernible on the tip, although it is shifted medially to 0.2 to 0.6 mm from the midline. There is a somewhat different distribution af papillae on the mid-region of the

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Fig. 2 Plots of locations of fungiform pap'llae on three rat tongues. Each dot represents one fuiigiform papilla. The tongue tip appears at the left and the intermolar eminence apFears at the right. Each papilla location is plotted by coordinate numbers obtained from tongue photographs. Units in millimeters of tongue surface.

tongue than on the tip. The greatest accumulation of papillae on the tin occurs at about 1 mm lateral to the midline and the distribution tails off regularly in a lateral direction. For the distribution of papillae exclusive of the paracentral line on the mid-region, the number of paDillae increases from the midline to about 2 mm laterally forming a nodal point and then decreases toward the lateral margin. The

number of papillae on the tip decreases from the midline to the lateral margin, while on the mid-region of the tongue the number of papillae reaches a maximum in the mid-lateral region and decreases in both directions, DISCUSSION

Rats in the current study had a mean of 187 9 papillae per tongue with a range

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Fig. 3 Plot of the mean numbers of fungiform papillae at various distances (anteriorposterior axis) from the origin of the coordinate system on the ventral tongue tip (squares, left ordinate). Cumulative percent of mean total papillae from anterior to posterior (circles, right ordinate). Tip Mmm)

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Fig. 4 Lateral distribution (medial-buccal axis) of mean numbers of fungiform papillae at various distances from the midline. On the right is the lateral distribution for the entire tongue. In the middle is the lateral distribution of papillae located within the anterior 4 mm of the tongues. The graph on the left shows the lateral distribution of papillae on the mid-region of the tongue from 5 to 20 mm posterior to the origin.

of 175 to 201 ( N = 10); while Fish et al. ('44) found 178.8 papillae per tongue with a range of 114 to 221 (N = 103). These results were not significantly different (t-test, p < 0.05), nor was there any difference found in the total number of papillae on the two lateral halves of the tongue in the current study. The 15% variability in the range of the total papillae per ani-

683

mal probably represents variation among animals rather than observational error since the procedure involved reproducible counting of macroscopic structures. An average of 51% of the total fungiform papillae are contained in an area of about 28 mmz on the first 4 mm of the tongue tip. This gives an average density of 3.4 papillae/mm2 on the tongue tip compared with only 1.3 papillae/mm2 in the mid-dorsal region from 4 mm to 17 mm posterior to the origin. In a recent unpublished electrophysiological experiment from this laboratory, the summated response of the whole chorda tympani nerve to stimulation of the tongue with NaCl was compared for stimulation of the tongue tip alone, the mid-dorsal region alone, and the entire tongue. The mid-dorsal region gave a response which was about 50% of the entire tongue response, while the tip gave a response of about 80% of the whole tongue. There were approximately equal numbers of papillae on the tongue tip and mid-region, but the higher density of papillae on the tongue tip produced a response which was about 30% greater than the response of the mid-region of the tongue. Lines of papillae are discernible in both the anterior-posterior and medial-lateral axes of the rat tongue. Mistretta ('72) reported that longitudinal (parasagittal) rows as well as diagonal (biased mediallateral) rows of fungiform papillae appear in the rat fetus. Mistretta agrees with Farbman ('65) that fungiform papillae first appear on the fetal rat tongue in a distribution which is similar to that on the adult tongue. Farbman points out that fungiform papillae begin to develop in the absence of nerve processes which he believes favors the idea that the stimulus for their development originates from the connective tissue. The lines of papillae on the tongue resemble the course of the main branches of the lingual nerve studied by Vij and Kanagasuntheram ('72) in the developing human tongue. The relationship between the distribution of fungiform papillae and the course of the fiber bundles which innervates them must be determined in the rat. The distribution of fungiform papillae is equivalent to the distribution of taste buds

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INGLIS J. MILLER, JR. AND ARTHUR J. PRESLAR

on the anterior rat tongue. Each rat fungiform papilla has a single apical taste bud (Fish et al., '44; Beidler and Smallman, '65) and no taste buds on the anterior tongue of the rat have been reported in the literature or observed in this laboratory which were not in association with fungiform papillae. Chorda tympani nerve fibers branch to innervate taste buds located in adjacent fungiform papillae; so the neural input from the fungiform taste buds is determined, in part, by their spatial distribution. Lateral enhancement and depression of the nerve response results from the neural interaction of adjacent papillae. Though the significance of spatial relationships among fungiform papillae to the process of chemoreception must be determined by mapping physiological preparations, this study has provided evidence of consistency in the number and distribution of papillae among a population of animals. ACKNOWLEDGMENTS

Patricia A. C . Ryan contributed technical assistance in the compilation of data and the preparation of illustrations. Daphne Styers provided clerical assistance in the preparation of the manuscript.

LITERATURE CITED Beidler, L. M., and R. L. Smallman 1965 R e newal of cells within taste buds. J. Cell Biol. 27 (2): 263-272. Farbman, A. I. 1965 Electron-microscope study of developing taste bud in r a t fungiform papilla. Dev. Biol., 11: 110-135. Fish, H. S., P. D. Malone and C. P. Richter 1944 The anatomy of the tongue of the domestic Norway rat. I. Anat. Rec., 89: 4 2 9 4 4 0 Miller, I. M. 1971 Peripheral interactions among single papilla inputs to gustatory nerve fibers. J. Gen. Physiol., 57 (1): 1-25. 1972 Integration of anion-cation inputs in rat chorda tympani fibers. In: Olfaction and Taste IV. D. Schneider, ed. Wissenschaftliche Verlagsgesellschaft MBH, Stuttgart, pp. 316 322. 1974 Branched chorda tympani neurons and interactions among taste receptors. 3. Comp. Neur., 158: 155-166. Mistretta, C. M. 1972 Topographical and histological study of the developing r a t tongue, palate and taste buds. Third Symposium om Oral Sensation and Perception: The Mouth of the Infant. J. F. Bosman, ed. Thomas, Springfield, pp. 163-187. Pfaffmann, C. 1970 Physiological and behavioural processes of the sense of taste. In: Ciba Foundation Symposium on Taste and Smell in Vertebrates. G. E. W. Wolstenholme and Julie Knight, eds. J. and A. Churchill, London,. pp. 31-50. Vij, S., and R. Kanagasuntheram 1972 Devel-. opment of the nerve supply to the human: tongue. Acta Anat., 81: 466-477.

Spatial distribution of rat fungiform papillae.

The objective of this study was to determine the spatial distribution of fungiform papillae on the rat's tongue. Since each fungiform papilla in the r...
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