Gary K. Beauchamp

Pseudogenization of a Sweet-Receptor Gene Accounts for Cats' Indifference toward Sugar

Although domestic cats (Felis silvestris catus) possess an otherwise functional sense of taste, they, unlike most mammals, do not prefer and may be unable to detect the sweetness of sugars. One possible explanation for this behavior is that cats lack the sensory system to taste sugars and therefore are indifferent to them. Drawing on work in mice, demonstrating that alleles of sweet-receptor genes predict low sugar intake, we examined the possibility that genes involved in the initial transduction of sweet perception might account for the indifference to sweet-tasting foods by cats. We characterized the sweet-receptor genes of domestic cats as well as those of other members of the Felidae family of obligate carnivores, tiger and cheetah. Because the mammalian sweet-taste receptor is formed by the dimerization of two proteins (T1R2 and T1R3; gene symbols Tas1r2 and Tas1r3), we identified and sequenced both genes in the cat by screening a feline genomic BAC library and by performing PCR with degenerate primers on cat genomic DNA. Gene expression was assessed by RT-PCR of taste tissue, in situ hybridization, and immunohistochemistry. The cat Tas1r3 gene shows high sequence similarity with functional Tas1r3 genes of other species. Message from Tas1r3 was detected by RT-PCR of taste tissue. In situ hybridization and immunohistochemical studies demonstrate that Tas1r3 is expressed, as expected, in taste buds. However, the cat Tas1r2 gene shows a 247-base pair microdeletion in exon 3 and stop codons in exons 4 and 6. There was no evidence of detectable mRNA from cat Tas1r2 by RT-PCR or in situ hybridization, and no evidence of protein expression by immunohistochemistry. Tas1r2 in tiger and cheetah and in six healthy adult domestic cats all show the similar deletion and stop codons. We conclude that cat Tas1r3 is an apparently functional and expressed receptor but that cat Tas1r2 is an unexpressed pseudogene. A functional sweet-taste receptor heteromer cannot form, and thus the cat lacks the receptor likely necessary for detection of sweet stimuli. This molecular change was very likely an important event in the evolution of the cats carnivorous behavior.

The vomeronasal organ: primary role in mouse chemosensory gender recognition.

Four experiments were conducted to determine the chemosensory modality that supports ultrasonic courtship vocalizations by male mice to females and to chemosignals from females. Both removal of the olfactory bulbs (Experiment 1) and removal of the vomeronasal organ (Experiments 2-4) produced similar deficits in the pattern of ultrasonic vocalizations elicited by conspecifics or their odors. Removal of the vomeronasal organ did not impair the ability to locate food buried under cage shavings. These results are consistent with the notion that the analysis of food related odors is subserved by olfaction and that vocalizations to sex chemosignals are elicited primarily by stimulation of the vomeronasal organ/accessory olfactory bulb. Removal of the vomeronasal organ did not induce seminal vesicle regression or lower plasma immunoreactive testosterone levels (Experiment 2) nor was an attempt to restore vocalizations with exogenous testosterone successful (Experiment 4). Thus the altered vocalization pattern following removal of the vomeronasal organ does not appear to arise as a motivational deficit mediated by androgens. Experiments 2 and 3 demonstrated that, in the absence of the vomeronasal organ, stimulation of other sensory systems can, to some extent, maintain the males tendency to vocalize more to females or their odors than to males or their odors. However, this responsiveness to females may rely upon additional behavioral cues. Previous experience also plays a considerable role in the response to chemosensory gender cues by males who lack their vomeronasal organs. Removal of the vomeronasal organ prior to adult heterosexual encounters (Experiment 3) virtually eliminated the males responsiveness to either anesthetized females or their chemosignals. Hence males require adult heterosexual experience with a functioning vomeronasal organ before other chemosensory systems acquire the ability to mediate gender recognition as measured by ultrasonic vocalizations.

Nutrient preference and diet-induced adiposity in C57BL/6ByJ and 129P3/J mice.

Purified carbohydrates and fats are usually palatable to humans and other animals, and their consumption often induces weight gain and accumulation of fat. In this study, we examined consumption of complex carbohydrates (cornstarch and Polycose) and fats (soybean oil and margarine) in mice from two inbred strains, C57BL/6ByJ and 129P3/J. At lower concentrations of liquid nutrients tested using two-bottle tests, when the amounts consumed had negligible energy content, the C57BL/6ByJ mice had higher acceptance of Polycose and soybean oil. This was probably due to strain differences in chemosensory perception of Polycose and oil. At higher concentrations, the mice consumed a substantial part of their daily energy from the macronutrient sources, however, there were no or only small strain differences in nutrient consumption. These small differences were probably due to strain variation in body size. The two strains also did not differ in chow intake. Despite similar energy intakes, access to the nutrients resulted in greater body weight (BW) gain in the C57BL/6ByJ mice than in the 129P3/J mice. The diet-induced weight gain was examined in detail in groups of 2-month-old C57BL/6ByJ and 129P3/J mice given ether chow, or chow and margarine to eat. Access to margarine did not increase total energy consumption of either strain. It increased BW and adiposity of the C57BL/6ByJ mice, but only after they reached the age of approximately 3 months. There were no differences in BW and adiposity between control and margarine-exposed 129P3/J mice. The results suggest that diet-induced adiposity in the B6 mice depends on age and does not depend on hyperphagia.

Flavor preferences in malnourished Mexican infants.

Taste and flavor preferences were evaluated in malnourished (n=113) and well nourished (n=87) Mexican infants tested between 2 and 24 months of age. Not all infants were tested in all conditions. The malnourished infants all suffered from second or third degree malnutrition (marasmus) and were tested 2-28 days after admission to a renutrition center. Well nourished control infants were tested at their own homes or neighbors homes. In the first experiment, testing consisted of brief presentations of moderate concentrations of sucrose (sweet), NaCl (salty), citric acid (sour) and urea (bitter). Infants were allowed to ingest as much of the taste substances as they wished. Volume of taste solutions and diluent was determined and provided the data for comparison between groups. In the second experiment, responses to casein hydrolysate and monosodium glutamate solutions were studied with soup serving as the diluent. The preferences of malnourished children for NaCl, citric acid and urea were not substantially different from those of the well nourished group. In sucrose tests, malnourished infants recently admitted to the renutrition center exhibited a decreased response. Malnourished infants ingested more casein hydrolysate and soup solution than soup alone. Control infants tended to respond in the opposite direction. All infants exhibited a preference for soup with MSG relative to plain soup.

Chemoinvestigatory and sexual behavior of male guinea pigs following vomeronasal organ removal

The vomeronasal organs of male guinea pigs were removed (VNX; n = 10) or males experienced sham surgery (Sham; n = 10). Subsequently a battery of chemosensory tests of investigatory responsiveness to conspecific urine was conducted. Additionally, male subjects were paired with female conspecifics for short and long periods and social and sexual behaviors were monitored. VNX males exhibited a depression in urine investigation and this depression became more profound following repeated testing and/or the passage of time. By 6.3 months following surgery, investigatory responsiveness to urine was practically eliminated. Maintenance of responsiveness to urine odors may require reinforcing input through the accessory olfactory system. In contrast to these effects on responsiveness to odors, VNX and Sham males were indistinguishable in their social and sexual behavior. These data indicate that male guinea pigs without a VNO: (1) Exhibit a depression of investigation of urine odors which is time dependent and which may involve an extinction-like process; (2) continue to discriminate classes of urine (e.g., urine from male vs urine from female conspecifics); and (3) exhibit normal sexual behavior. The vomeronasal organ in the male domestic guinea pig is apparently critical for the maintenance of normal responsiveness to sex odors but, in its absence, other sensory systems are capable of maintaining normal sexual behavior under conditions of laboratory testing.

Individual odortypes: interaction of MHC and background genes

Genes of the major histocompatibility complex (MHC) influence the urinary odors of mice. Behavioral studies have shown (1) that mice differing only at MHC have distinct urinary odors, suggesting an MHC odor phenotype or odortype; (2) that the MHC odortype can be recognized across different background strains; and (3) that the MHC odortype is not an additive trait. Very little is known about the odorants underlying this behavioral phenotype. We compared urinary volatile profiles of two MHC haplotypes (H2b and H2k) and their heterozygous cross (H2b×H2k) for two different background strains (C57BL/6J and BALB/c) using solid phase micro-extraction (SPME) headspace analysis and gas chromatography/mass spectrometry (GC/MS). Both MHC and background genes substantially influence the volatile profile. Of 148 compounds screened, 108 of them significantly differ between the six genotypes. Surprisingly, for numerous compounds, their MHC associations are moderated by background genes (i.e., there is a significant MHC × background interaction effect in the statistical model relating genotype to relative compound concentration). These interactions account for nearly 30% of the total genetic effect on the volatile profile. MHC heterozygosity further extends the odortype diversity. For many compounds, the volatile expression for the heterozygote is more extreme than the expression for either homozygote, suggesting a heterozygous-specific odortype. The remarkable breadth of effects of MHC variation on concentrations of metabolites and the interaction between MHC and other genetic variation implies the existence of as yet unknown processes by which variation in MHC genes gives rise to variation in volatile molecules in body fluids.

In search of the chemical basis for MHC odourtypes

Mice can discriminate between chemosignals of individuals based solely on genetic differences confined to the major histocompatibility complex (MHC). Two different sets of compounds have been suggested: volatile compounds and non-volatile peptides. Here, we focus on volatiles and review a number of publications that have identified MHC-regulated compounds in inbred laboratory mice. Surprisingly, there is little agreement among different studies as to the identity of these compounds. One recent approach to specifying MHC-regulated compounds is to study volatile urinary profiles in mouse strains with varying MHC types, genetic backgrounds and different diets. An unexpected finding from these studies is that the concentrations of numerous compounds are influenced by interactions among these variables. As a result, only a few compounds can be identified that are consistently regulated by MHC variation alone. Nevertheless, since trained animals are readily able to discriminate the MHC differences, it is apparent that chemical studies are somehow missing important information underlying mouse recognition of MHC odourtypes. To make progress in this area, we propose a focus on the search for behaviourally relevant odourants rather than a random search for volatiles that are regulated by MHC variation. Furthermore, there is a need to consider a ‘combinatorial odour recognition’ code whereby patterns of volatile metabolites (the basis for odours) specify MHC odourtypes.

Cats Lack a Sweet Taste Receptor

Domestic cats (Felis silvestris catus) (herein referred to as “cats”) are neither attracted to, nor show avoidance of the taste of sweet carbohydrates and high-intensity sweeteners (1-3), yet they do show a preference for selected amino acids (4), and avoid stimuli that taste either bitter or very sour to humans (1,4). Consistent with this behavioral evidence, recordings from cat taste nerve fibers and from units of the geniculate ganglion innervating taste cells demonstrated responses to salty, sour, and bitter stimuli as well as to amino acids and nucleotides, but showed no response to sucrose and several other sugars (4-11). The sense of taste in cats appears similar to that of other mammals with the exception of an inability to taste sweet stimuli. n nBecause only the sweet taste modality appears absent, we postulated that the defect in cats (and likely in other obligate carnivores of Felidae) lay at the receptor step, subtending this modality. The possible defects at the molecular level could range from a single to a few amino acid substitutions, such as is found between sweet “taster” and “nontaster” strains of mice (12-14), to more radical mechanisms, such as an unexpressed pseudogene. n nTo distinguish among these possibilities, we identified the DNA sequences and examined the structures of the 2 known genes Tas1r2 and Tas1r3 that encode the sweet taste receptor heteromer T1R2/T1R3 in other mammals. We compared these with the sequence and structure of the same genes in dogs, humans, mice and rats, all species that respond to sweet stimuli.

Chemical Communication in the Guinea Pig: Urinary Components of Low Volatility and Their Access to the Vomeronasal Organ

Guinea pigs (Cavia sp.) utilize chemical information from conspecifics in the regulation of social behavior (Beauchamp et al., 1977; King, 1956; Rood, 1972), We have been Investigating a number of aspects of this communication system. Here, we will present observations and evidence from behavioral and chemical studies that suggest that information transfer among individuals is mediated at least in part by compounds of low volatility. If compounds of low volatility are used, the receptor organ(s) and the means by which the compounds reach the organ(s) are called into question. Volatile compounds can easily reach the olfactory receptor cells during inspiration; non-volatile substances presumably must reach appropriate receptors by some other means. We have hypothesized that the vomeronasal organ is involved in responses to chemical signals, particularly those of low volatility. We also will describe studies investigating access of non-volatile material to the vomeronasal organ.

Chemical and Behavioral Complexity in Mammalian Chemical Communication Systems: Guinea Pigs (Cavia Porcellus), Marmosets (Saguinus Fuscicollis) and Humans (Homo Sapiens)

The odorous secretions produced by mammals have long interested man. In some ancient civilizations, the scents produced by sacred or powerful animals were worn by men in an effort to capture some of its power (Kingston, 1965). In more recent times, organic chemists and perfumers have sought to identify and employ the volatile constituents of mammalian secretions and excretions in perfumes and colognes while animal behaviorists and reproductive biologists have explored the role such constituents play in olfactory chemical communication. With the establishment that volatiles in excreted body fluids play a significant role in mammalian communication organic chemists have become interested in the isolation and identification of these compounds. Such studies require an interdisciplinary approach.