Biology of Taste Buds and the Clinical Problem of Taste Loss

Transcription

1 70 THE ANATOMICAL RECORD (NEW ANAT.) FEATURE ARTICLE Biology of Taste Buds and the Clinical Problem of Taste Loss GINA M. NELSON Taste buds are the anatomical structures that mediate the sense of taste. They comprise taste cells and nerve fibers within specialized epithelial structures. Taste cells are traditionally described by histologic methods as basal, dark, intermediate, and light cells, with the nerve fibers surrounding and infiltrating the taste buds. By means of immunohistochemical methods, taste cells and gustatory nerve fibers can be classified in functional groups based on the expression of various cell adhesion molecules and other proteins. When taste buds become damaged, the loss of the ability to taste results. This loss is not uncommon and can impact health and quality of life. Patients who receive radiation therapy for head and neck cancer often experience taste loss, which leads to compromised nutritional intake and a worse outcome than patients who do not experience taste loss. The mode of radiation damage to taste cells and nerve fibers has been investigated using cell adhesion molecules, synaptic vesicle proteins, and other cell markers. The light and intermediate cells are preferentially affected by ionizing radiation, whereas the nerve fibers remain structurally intact. Experimental studies of radiation-induced taste loss are performed via a unique animal/human model. Anat. Rec. (New Anat.) 253:70 78, Wiley-Liss, Inc. KEY WORDS: taste buds; taste loss; head/neck cancer; radiation therapy; radiation model; cancer therapy WHAT MAKES UP THE SENSE OF TASTE? Of the five senses, taste is the sense which is least thought about by most people. Unlike the senses of vision and hearing, taste does not have much impact on our day-to-day life or does it? Taste buds play a greater role in our lives than we might imagine. In combination with our sense of smell, we are able to enjoy the aroma and taste of our food and drink. More importantly, we recognize spoiled food, if not by the smell then by the terrible taste. For our ancestors more than for modern humans, taste warned of poisonous Dr. Nelson has long had an interest in chemoreceptive sciences, beginning with invertebrate chemoreception. She has an MD and PhD from the University of Colorado Health Sciences Center, where she developed an animal model of radiation-induced taste loss. Dr. Nelson is currently a resident in anatomic pathology at the University of Iowa Hospitals and Clinics and plans to continue work on the animal model and human studies in taste loss through the Department of Anatomy and Cell Biology after residency Wiley-Liss, Inc. plants, as they have a very bitter taste due to the presence of various alkaloid compounds. What happens when our sense of taste goes awry? The understanding of the mechanisms of vision and hearing is far ahead of the understanding of the mechanisms of taste. Glasses and artificial lenses are available to correct vision deficits, and hearing aids are available for those with diminished hearing, but what corrective aids are available for the loss of taste? Have you ever known a person, a relative or a friend perhaps, who cannot taste? What is the world of taste like for that person? What causes taste loss? Is the ability to taste affected by illness or by therapies designed to treat these illnesses? WHERE ARE MY TASTE BUDS? Taste buds are the anatomical structures which contain the receptor cells that mediate the sense of taste. Taste buds are found in the oral cavity, primarily on the tongue but also on the palate, back of the mouth, pharynx, epiglottis, and larynx. The tongue (Fig. 1) is covered with numerous papillae which come in four varieties. Filliform papillae are the most numerous and appear as short, rough structures covered with thick keratinized epithelium (they feel particularly rough on a cat s tongue). They do not contain taste buds. The fungiform papillae are dispersed across the surface of the tongue. They are more box-like, with a connective tissue core and a thin covering of epithelium. Most of the fungiform papillae contain a single taste bud on the tip. The larger circumvallate papillae are located on the posterior aspect of the dorsal surface, appearing as pincushions with a surrounding trough, called a crypt. The crypt is lined by an epithelium, called the gustatory epithelium, which contains several taste buds. The pores of the taste buds open into the crypt. In humans there are circumvallate papillae, but in rodents (as in mice or rats) there is only one. Foliate papillae lie on the lateral sides of the tongue and appear like slits. These are less developed in rats and humans than they are in other species. Each taste bud is oval (Fig. 2) and opens to the epithelial surface via a small opening called a taste pore. From

2 REVIEW THE ANATOMICAL RECORD (NEW ANAT.) 71 Fig. 1. Dorsal surface of tongue. Pictured is the upper surface of the tongue showing the location of the pincushion shaped circumvallate papillae on the very back of the tongue and the fungiform papillae over the surface between the circumvallate papillae and the tip of the tongue. this pore protrudes the microvilli arising from the tips of the individual taste cells. Each taste bud contains taste receptor cells and support cells. Taste cells are described as basal, dark, intermediate, and light, based on electron microscopic characteristics 1 (Fig. 2). The basal cells are at the base of the taste bud and constitute a proliferative population of cells. They divide to produce postmitotic light, intermediate, and dark taste cells with a life span of days. Dark cells are defined by a dark cytoplasm (electron-dense), dense-core granules (small vesicles with a dark center) at the tip of the cell, indentations in the nuclear membrane, and collections of heterochromatin (the DNA) along the inner edge of the nucleus. Light cells are characterized by a light cytoplasm (electron-lucent), clear vesicles and mitochondria in the tip of the cell, and a round to oval nucleus with less heterochromatin (DNA) along the inner edge. Intermediate cells have characteristics that are intermediate between the light *Authors vary in their use of the different systems of nomenclature. While some correlations have been made between the two systems, they are not totally equivalent. The information here is given in relation to the article in which it was originally presented, maintaining the system by which it was described. and dark cells. Which taste cell type is actually the receptor has not been proven, although every type has been proposed. Taste cells have also been designated as Type I, II, III, and IV, with Type I being similar to dark cells, Type II similar to light cells, and Type IV being the basal cells. In this schema, the Type III cells are like intermediate cells and are thought to be the gustatory receptor cells, 2 primarily due to the presence of what are believed to be afferent synaptic contacts* (afferent nerve fibers carrying information to the brain). The dense-core vesicles are located in dark cells and to a lesser extent in intermediate cells 1 in association with both the tip of the cells and the presynaptic regions. These vesicles are thought to contain serotonin. 3 Fungiform taste buds are innervated by the chorda tympani branch of cranial nerve VII, while the circumvallate taste buds are innervated by cranial nerve IX. Taste buds in the pharynx are innervated by cranial nerve X. In the region of the taste buds, nerve fibers are described by anatomical location (refer to Fig. 2). The subepithelial plexus are the nerve fibers in the connective tissue beneath the gustatory epithelium (the epithelium containing the taste buds). The basal plexus nerve fibers are in the basal portion of each bud, forming a kind of a nest around the basal cells. Intragemmal fibers are those within the taste bud, and perigemmal fibers are those around the taste bud. The intragemmal nerve fibers are the nerve fibers which receive synaptic contacts from all three types of taste cells. 4 While the perigemmal fibers are generally thought to be outside of the taste bud, some perigemmal nerve fibers enter the taste bud from the side. Some perigemmal nerve fibers reach the surface of the epithelium; others reach around the taste pore. In addition, both Type II and III cells form two types of synaptic contacts with the intragemmal nerve fibers, thought to represent efferent (efferent nerve fibers carrying information from the brain) and afferent synaptic contacts. 5 It has been proposed that the substance P containing fibers within the taste bud may mediate oral pain. 6 A nutritional, or trophic, interaction (i.e., one cell emits a substance that a second cell needs to grow) between the nerve fiber and taste buds exists. Interruption of the nerve fiber by either cut or crush injury results in the Taste buds play a greater role in our lives than we might imagine. disappearance of the taste buds (Fig. 3). If the nerve fiber is allowed to grow back, the taste buds will reappear. This indicates that there is an unknown molecule originating from the nerve fiber that is required for the taste buds to retain their form. The mechanisms of sensory transduction of various taste stimuli across the taste cell membrane is a topic worthy of its own review, and the reader is referred to excellent papers on this topic. 7 ANATOMICAL CONCEPTS BASED ON CELL MARKERS Until recently, the elements of the taste system have been described anatomically and histologically. An alternative method is to describe the components of the taste system based on their function, which makes sense biologically. By comparison, vision receptors are the rods and cones, and the vestibular and cochlear nerves transmit specific sensory information in the ear.

3 72 THE ANATOMICAL RECORD (NEW ANAT.) REVIEW Fig. 2. Schematic of a normal taste bud. All of the elements of a taste bud are illustrated. The taste bud contains basal cells (B) in the lower portion of the bud and all three types of taste cells extending upward to the opening at the top, the taste pore. The light cells (L), intermediate cells (I), and dark cells (D) are pictured with some of the features that characterize each cell type. In addition, the intragemmal and perigemmal nerve fibers are seen. The nerve fibers are present in the subepithelial connective tissue and enter each bud from the base. The red fibers represent the nerve fibers which contain synaptic vesicle proteins, and the black fibers represent nerve fibers which contain peptides. S, synapse. Fig. 3. Damage to the taste bud following nerve injury. After crushing or cutting the nerve (e.g., glossopharyngeal) that innervates the taste buds, the taste buds degenerate. Small, atrophic buds remain, as seen in the illustration. If the nerve fiber regenerates, the taste buds will grow back.

4 REVIEW THE ANATOMICAL RECORD (NEW ANAT.) 73 Fig. 4. The technique of immunohistochemistry. The protein of interest (pentagon in the illustration), termed the antigen, is isolated and purified and then injected into the host mouse. The mouse s immune system sees the antigen as a foreign molecule and makes antibodies against it. The antibodies are then taken from the mouse s serum. A thin piece of tissue from a second animal is put on a glass slide, and the antibody is applied to it. The antibody will recognize the antigen in the tissue (the pentagon) and stick to it. In order to visualize where the antibody is located, a secondary antibody is applied which recognizes the first. The secondary antibody has a tag on the end (star) which can be seen in a microscope equipped for fluorescence microscopy. These descriptors are related to the function of the anatomical structure. It would be logical to take the same approach for the sense of taste and designate a salt receptor or a bittertaste fiber. However, the details are not known at a sufficient level to always determine the functional designations. Investigators in many laboratories are improving on the knowledge of the taste system, so this descriptive system may be realized in the future. One way to approach this is to identify groups of taste cells or nerve fibers based on the expression of a particular protein. With a technique called immunohistochemistry, an antibody is made which recognizes a particular protein for example, protein X (Fig. 4). Simply, an amount of protein X is injected into a mouse, the host animal. The immune system of the host generates antibodies directed against protein X. The serum is removed from the animal, and the new anti-x antibodies are purified. Thin sections of tissue from the animal being investigated (usually not a mouse) are put on slides, and the anti-x antibodies are added. The antibodies attach to the X molecules in the tissue. Then a secondary antibody with a fluorescent tag is attached to the first antibody. A microscope equipped to view the fluorescent tag is then used to visualize where the anti-x antibodies attached. The correlation can be made that protein X resides in the location where fluorescent patterns are seen. The markers used to identify cells and nerve fibers can be cell surface molecules, neurotransmitters, structural proteins, synaptic vesicle proteins, peptides, blood group markers, enzymes, lectins, or many other types of proteins. The identification of a group of taste cells or nerve fibers that share a given histochemical property suggests that these cells or nerve fibers have a common biological characteristic which may be more closely related to a common function. Taste cells or nerve fibers classified in this manner can be studied in various experimental situations. Often the expression of a particular protein is found primarily in a histologically designated type of taste cell (i.e., light cells), although only a few of the light cells contain the protein. Sometimes a few cells from two groups will express a particular protein (i.e., a few of the light and intermediate cells). This suggests that the histological classifications of light, intermediate, and dark do not correlate with function. Examples of various proteins expressed by taste cells include the blood group antigens, 8 the transmembrane G protein gusducin 9 neural cell adhesion molecule (NCAM), 10 the calcium binding protein calbindin, 11 and keratins. 12 Some are located in specific taste cell types; others are not. Many other examples are described in the literature. Using light and electron microscopic immunohistochemistry, we demonstrated the presence of NCAM 10 and a form of growth associated protein (GAP) in taste cells recognized by the antibody designated B50 13 (Fig. 5). NCAM appears as smooth, continuous outlines on long, thin, distinct taste cells identified as light or intermediate cells. The B50 antibody produces a diffuse label throughout taste cells identified as intermediate or dark cells, located in the mid to apical portion of the taste bud. These cells appear different from the thin, elongated cells labeled by the NCAM antibody. Not all of the intermediate and dark cells label. Experiments with NCAM and B50 together indicate that these proteins occur on separate populations of taste cells. No cell in normal taste buds ever showed reactivity to both of these antibodies together. ANATOMICAL CONCEPTS BASED ON NERVE FIBER MARKERS Nerve fibers can also be described in a similar fashion. In the construct of a sensory system, there are nerve fibers to bring the signal of the perceived stimulus to the brain (the nerve fibers which are postsynaptic to the taste cells, the afferent fibers) and nerve fibers to bring modulating information to the sensory cells (efferent fibers). There may also be nerve fibers

5 74 THE ANATOMICAL RECORD (NEW ANAT.) REVIEW Fig. 5. Taste bud with antibody-labeled taste cells. After application of an antibody which recognizes NCAM, some of the light to intermediate taste cells appear bright, having a long, thin outline (taste cells with white dots in illustration). The application of the B50 antibody labels some of the dark cells with a more diffuse pattern (bright taste cell without dot). In this example, the nerve fibers are not labeled. Fig. 6. The effects of radiation on taste buds. At the peak of radiation damage (approximately 7 days), some of the light to intermediate taste cells are degenerating, but the dark cells and all subtypes of nerve fibers remain intact. The nerve fibers containing synaptic vesicle proteins (red) and the nerve fibers containing peptides (black) remain as they were in the normal taste bud. The overall pattern of degeneration is different than that seen in the taste buds affected by nerve cut (refer to Fig. 3).

6 REVIEW THE ANATOMICAL RECORD (NEW ANAT.) 75 to carry visceral information (hot, cold, pain, etc.) which may or may not be located within the taste bud. In an effort to identify these various types of nerve fibers, much like the taste cells, immunohistochemical techniques were utilized to demonstrate that different nerve fibers express different proteins. For example, many perigemmal fibers contain the peptide substance P, while others contain calcitonin gene-related peptide (CGRP). The intragemmal nerve fibers contain synaptic vesicle proteins, like synaptophysin. 14 (Of note, even though all of the types of taste cells, light, intermediate, and dark, make synaptic contacts with the intragemmal nerve fibers, 4 none of the synaptic vesicle proteins to date have been found to label the presynaptic vesicles located within the taste cells). Numerous basal plexus nerve fibers and nerve fibers in the dermis or in the core of the fungiform papillae also contain the synaptic vesicle proteins and peptides. Electron micrographs show that small vesicles within these nerve processes, measuring nm in diameter, are the same size as the vesicles described as containing the synaptic vesicle proteins in other locations. The nerve fibers that label with synaptic vesicle proteins are postsynaptic to some taste cells. 14 All of the nerve fibers, both intragemmal and perigemmal, can be identified with proteins common to most neurons, like protein gene product 9.5 (PGP 9.5) and S When the synaptic vesicle proteincontaining nerve fibers are compared with those containing peptides or PGP 9.5 using double labeled fluorescence immunohistochemistry, the two types of nerve fibers are not distinct groups, and the location of the proteins does not correlate exactly with the anatomical classifications. The nerve fibers containing synaptophysin have a small subset that also contain CGRP. There are occasional intragemmal nerve fibers that contain CGRP only. Most perigemmal nerve fibers show labeling with either synaptophysin or CGRP. Analysis of SV2 (another synaptic vesicle protein 16 ) and CGRP gives slightly different results. For both the intragemmal and perigemmal nerve fibers, there is a group of fibers that contain both proteins, and there is a group of fibers that contain SV2 only and CGRP only. All synaptophysin nerve fibers are also PGP 9.5 nerve fibers, but the reverse is not true. Even though there is some overlap in the described categories of gustatory nerve fibers, the results of examining the distribution of these proteins in the gustatory nerve fibers allows their division into functional groups: postsynaptic intragemmal (labeled with synaptic vesicle proteins), nonpostsynaptic intragemmal (labeled with peptides), perigemmal (labeled with peptides), and nerve fibers around the taste pore (labeled with SV2). This gives us a tool to follow what happens to these categories of nerve fibers during experimental manipulation, as described below. TASTE LOSS IN HUMANS While not discussed widely in the biochemical literature, ageusia (taste loss) and hypogeusia (decrease in taste) and dysgeusia (abnormal taste) are widespread and associated with a variety of illnesses, from common to obscure. Taste loss occurs as a natural phenomenon of aging and also in response to normal changes such as pregnancy and menopause. 17 Poor dentition and hygiene are common oral conditions that affect taste. Patients with xerostomia (dry mouth), Sjögren syndrome (inflammation of the salivary glands resulting in a dry mouth), and zinc deficiency may also experience taste loss. Other conditions in which taste loss may occur include liver and kidney disorders, diabetes mellitus, depression, and surgical procedures around the chorda tympani or glossopharyngeal nerve. Patients with head trauma and epilepsy may also experience taste loss. Taste loss can range from mild to severe, resulting in subsequent decrease in nutritional intake. (Many investigators contributed to the accumulation of this knowledge. The reader is referred to the many chapters that cover these topics in more detail in Getchell et al. 18 ) Various types of therapy can also induce taste loss. Numerous drugs are associated with taste loss. However, an incomplete understanding of how the taste system works and the interaction of drug compounds with the taste system make it difficult to assign the taste loss to a drug alone. One cannot rule out that the taste loss is caused by the underlying disease process. Most of the drugs associated with taste loss affect the turnover of cells, as is seen in other systems, but other mechanisms are possible. Drugs usually induce a temporary effect which diminishes after the drug is discontinued. Chemotherapy employs drugs associated with taste loss. A few examples include methotrexate and dexamethasone, antihypertensives, antimicrobial agents, and antiproliferative agents. 18 RADIATION THERAPY INDUCED TASTE LOSS Radiation is often used either alone or in addition to surgery to treat various types of cancers. The typical dose for patients with head and neck cancer or oral cancer is 5,000 7,000 cgy. (A centigrey (cgy) is the deposition of 1 erg of energy per 100 g of tissue.) It is administered in divided doses of about 180 cgy/week until the desired dose is achieved. Radiation is composed of charged particles that disrupt the electron orbital structure of the atoms in the tissue, causing tissue destruction. 19 The theory behind the therapy is to disrupt the proliferative capacity of the tumor, thus destroying it, while doing as little damage as possible to the normal tissue. Since the tissue lining the mouth and the gastrointestinal tract divides at a faster rate than tissues of other organs (e.g., liver), they are more susceptible to radiation damage. The resulting side effects to the oral cavity include mucositis (swelling and tenderness of the oral mucosa with sloughing off of dead cells), xerostomia, and taste loss. Of particular interest is the taste loss that occurs with the administration of radiation therapy, termed postirradiation gustatory dysfunction. It occurs following administration of radiation to the region of the oral cavity and thus the taste buds. In some patients, the taste loss can be severe. The loss of taste has been reported in the literature only as case reports and as occasional small studies. The loss of taste due to radiation therapy is a common problem which is underrepresented in the literature. In patients who lose their sense of taste, one result is a marked decrease in the ability to eat and thus a decrease in nutrition

7 76 THE ANATOMICAL RECORD (NEW ANAT.) REVIEW intake. These patients experience greater weight loss than those patients who do not report a change in taste, 20 and it has been well documented that these patients have a worse outcome than the patients who do not lose their sense of taste and are able to maintain their food intake and nutritional support. 21 Nutritional supplements have been shown to positively impact cancer therapy when administered in addition to the therapy for the disease. 22 The changes in the taste thresholds for all tastes (sweet, sour, bitter, salty) varies among treated patients. Quinine taste (bitter) is most consistently lost, but the loss of sugar and salt taste varies considerably among patients. Decreases in taste thresholds begin from treatment with as little as cgy. 23 Also, a discrepancy is noted between the speed with which taste sensation is lost and the deterioration of the taste cells. The taste buds degenerate 6 7 days after irradiation, but taste alteration is seen as early as 2 3 days after irradiation. In mice, following a single radiation dose between 1,000 and 4,000 cgy, taste buds are either entirely destroyed or, if they remain, lose 30 50% of their cells. The taste bud degeneration peaks at 9 days after injury, and then the taste buds begin to regenerate. 24 In another study, 25 single doses of 850 cgy cause a smaller number of taste buds to degenerate, and recovery begins sooner. At a single dose of 2,200 cgy, the taste buds degenerate much faster. This illustrates that the number of taste buds that degenerate is related to the size of the dose. When permanent damage occurs, it is usually with accumulated doses exceeding 6,000 cgy. Long-term effects include lowered taste detection and threshold levels as well as xerostomia. The maximum tolerance dose giving a 50% complication rate is approximately 4,000 6,500 cgy for xerostomia and 5,000 6,500 cgy for taste loss. 26 The loss of taste does not vary by type of radiation (i.e. neutron vs. photon radiation). 27 Radiation affects elements of cells that cannot easily be repaired or replaced, namely the DNA. As a consequence, proliferative cells are most sensitive to the effects of radiation. Inside the taste buds are the nondividing nerve fibers and a proliferative population of taste cells. This suggests two possible sites of radiation damage leading to taste loss: (1) the intragemmal nerve fibers and (2) the taste cells. If the nerve fibers are the site of damage, one possibility could be a significant physical loss in the population of postsynaptic intragemmal nerve fibers. Since the neurons are a nondividing population of cells, they are thought to be generally radioresistant. However, disruption of the functional integrity of the neuron could lead to the symptom of taste loss. Synaptic uncoupling or disruption of membrane integrity leading to a disruption in the contact between the taste cells and nerve fibers, resulting in the inability to conduct action potentials, is a possibility. A similar finding was demonstrated in the disruption of neuromuscular junctions in mouse tongues following a single dose of radiation. 28 Other investigators have actually proposed the nerve fiber as the site of damage leading to taste loss but mostly incidentally. 24,25 Radiation affects elements of cells that cannot easily be repaired or replaced, namely the DNA. The second possible site of damage is the taste cells. Previously it has been demonstrated that following irradiation the cells within the taste bud lose their characteristic histological appearances (light, intermediate, and dark) and all appear as intermediate cells. 25 The disruption of the proliferative capacity of the taste cells would cause stem cells to stop dividing, and, once the current receptors die off, no new ones would be there to replace them. This would be experienced as a loss of taste. It does not, however, account for the changes in taste measured at 2 3 days following irradiation in both animals and humans. This is more likely to be a disruption of the current receptor cells, possibly via membrane damage causing loss of structural integrity, or loss of the synaptic contacts. Other possibilities suggested include radiation-induced changes in metabolism, possibly associated to depleted zinc stores. 20 In an attempt to test these various hypotheses, the projects described herein use a novel approach of combining histological evaluation with behavioral assessment in an effective model of radiation-induced taste loss in rats, as well as extending the studies to human cancer patients receiving radiation therapy. In this model, hypotheses about the function of taste cells and nerve fibers can be formulated based on quantitative behavioral data. The tool we have designed to direct the radiation to the surface of the tongue results in a method of radiation which eliminates the problematic side effects encountered with conventional radiation, typically mucositis and xerostomia. In addition, the labels for the various subpopulations of taste cells and nerve fibers make it possible to follow what happens to each of these components of the taste system following radiation. The eventual goal of this model is to understand the biological mechanisms underlying radiation-induced taste loss so that methods to prevent taste loss can be developed and thus improve the quality of life and treatment outcome of these cancer patients. RADIATION-INDUCED TASTE LOSS IN ANIMALS In studies described previously, 29 the effects of radiation on the taste system were examined in rats given a single dose (1,700 cgy) of radiation to the oral cavity. Behavioral measurements were made based on the consumption of either a 1.8% NaCl (salt) solution or a quinine-hcl (bitter) solution and correlated with the histological changes in two groups of nerve fibers and two groups of taste cells at various time points after radiation. The nerve fibers followed were labeled with either synaptophysin or CGRP, and the taste cells followed were labeled with either NCAM antibodies or B50. Briefly, following irradiation, two changes in the consumption of 1.8% NaCl are noted: (1) a decrease in the total volume of fluid consumed and (2) an increase in the amount of NaCl consumed. Both of these changes begin to occur on day 4 (day 0 is the day the animals received radiation), peak

8 REVIEW THE ANATOMICAL RECORD (NEW ANAT.) 77 at days 7 8, and return to preirradiation levels by day 11. The statistical analysis showed no significant difference prior to radiation between the two groups of animals (experimental and control). Following irradiation, a significant difference was seen for the NaCl consumption on days 7 and 8. Following the dose of radiation, the distribution of the nerve fibers labeled with synaptophysin or CGRP does not change. However, the NCAM-labeled cells show a dramatic change in appearance. At 6 days, the NCAM cells appear normal, with one or possibly two cells demonstrating label only in small patches. At day 7 there is a dramatic change. Only rare, abnormalappearing cells labeled with NCAM remain. Also, the rare cells with any NCAM are also immunoreactive with the GAP antibody B50. The normal pattern of NCAM labeling begins to return at 11 days, and NCAM-labeled cells are more numerous by 16 days. The change in the pattern of labeling for antibody B50 is different from that observed for NCAM. Unlike NCAM antibody labeling, B50 labeling retains its normal pattern until 11 days. At this time, the B50-labeled cells begin to appear irregular in shape and remain so throughout 21 days. Again, the two proteins begin to appear in the same taste cells at day 7. These findings, which are summarized in Figure 6, indicate that changes in taste acuity do indeed occur and can be measured effectively and correlated with histological changes. The loss of the NCAM-labeled taste cells correlates with the noted changes in consumption of 1.8% NaCl. However, the B50-labeled cells remains intact. This suggests that NCAM-labeled cells could be NaCl receptors, while the B50-labeled cells are not. Clearly the division of these two groups of taste cells is altered by the radiation when the two proteins begin to colocalize. The biology of the cells is altered by the radiation, but its meaning is not yet understood. CAN TASTE LOSS BE TREATED? Two questions arise naturally out of these findings. Can the same type of behavioral and histological measurements be done in humans? And how will this information impact taste loss in human cancer patients? Psychophysical measurements of taste acuity and taste thresholds have been reviewed extensively in the literature (see Bartoshuk 30 ) Detection thresholds are stable over time (M. Linschoten, personal communication) and can be measured using a two alternative forced choice procedure. 31 The premise is to offer the subject two solutions (one water, one containing a tastant) to taste, and the subject chooses which one has a taste different from water. When the patient cannot distinguish between correct and incorrect for a given tastant concentration, the patient is believed to be at his or her threshold for that tastant. As patients receive radiation or chemotherapy, the changes that occur in the taste thresholds of any or all of the four tastants (sweet, sour, bitter, salty) can be followed. It is even possible to monitor the recovery, or lack thereof, of taste thresholds in these patients over long periods of time. The purpose of this entire area of study is twofold: to improve on the quality of life of cancer patients receiving radiation therapy and to improve the outcome of therapy by maximizing the patients ability to eat and maintain nutritional support. Once the biology of the taste loss is understood including the degree of taste loss and what taste qualities are lost modifications to the way the patient receives treatment can be made. One possibility is to design a diet that maximizes on the remaining abilities to taste, resulting in the most palatable diet available to the patient. This would require individual diet management, depending on the thresholds that change for that patient. This is similar to other specialized diets, such as diabetic diets. Another approach would be to protect the taste cells from the radiation damage. A fine balance has to be made between using enough radiation to kill the tumor cells and keeping damage to normal tissue at a tolerable level. A local application of a molecular substance in a foam or gel vehicle applied to the oral mucosa and tongue to impart a temporary resistance to the penetrating photons may reduce the damage to taste cells. Such a compound is only theoretical at the present. In summary, radiation-induced taste loss is a real problem that has a significant effect on the treatment of cancer. With information gathered from the animal model and human studies, we hope to achieve a better understanding of the function of the taste system and to positively impact the treatment outcome of cancer patients receiving radiation therapy. ACKNOWLEDGMENTS I would like to thank Drs. Mary J.C. Hendrix, Richard Lynch, and Thomas E. Finger for their support and encouragement. Also thanks to Jolene Redvale for review of the manuscript and especially Ken Nelson for the invaluable assistance with the computer graphics. The clip art in Figures 1 and 4 was provided by the Corel Corporation. LITERATURE CITED 1 Kinnamon JC, Taylor BJ, Delay RJ, Roper SD (1985) Ultrastructure of mouse vallate taste buds. I. Taste cells and their associated synapses. J Comp Neurol 235: Kanazawa H, Yoshie S (1996) The taste bud and its innervation in the rat as studied by immunohistochemistry for PGP 9.5. Arch Histol Cytol 59: Fugimoto S, Ueda H, Kagawa H (1987) Immunocytochemistry of the localization of 5-hydroxytryptamine in monkey and rabbit taste buds. Acta Anat 128: Royer, SM, Kinnamon JC (1988) Ultrastructure of mouse foliate taste buds: Synaptic and nonsynaptic interactions between taste cells and nerve fibers. 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