0300-9629/92$5.00 + 0.00
Camp. Eiochem. Physiol. Vol. IOIA, No. I, pp. 97-102, 1992 Primed in Great Britain
0
1991 Pergamon
Press plc
TASTE SYNERGISM BETWEEN MONOSODIUM GLUTAMATE AND 5’-RIBONUCLEOTIDE IN MICE Yuzo
NINOMIYA, SHUJI KURENUMA,* TAKAYUKI NOMURA, HAJIMU UEBAYASHI and HIROSADA KAWAMURA
Department
of Oral Physiology and *Oral Surgery, Asahi University School of Dentistry, Hozumi, Motosu, Gifu 501-02, Japan. Telephone: 05832-6-613 1 (Received
17 April
1991)
Abstract-l. Strain differences of mice were found in the taste synergism between monosodium L-glutamate (MSG) and disodium 5’-guanylate (GMP). 2. Magnitudes of chorda tympani responses to the mixture of MSG and GMP over the sum of responses to each component were greater in the order of C3H/HeSlc(C3H) > C57BL/6CrSlc(C57BL) > BALB/cCrSlc(BALB) mice. The greatest synergism was observed in response to the mixture of 0.03 M MSG and 0.1 mM GMP, to which responses were about 2.6, 1.8 and 1.4 times greater than the sum of each component in C3H, C57BL and BALB mice, respectively. 3. Magnitudes of inhibition of MSG and mixture responses by the lingual treatment of proteolytic enzyme, Pronase E, were greater in the same order of C3H > C57BL > BALB mice as that observed in magnitudes of the synergism. These results suggest that there exists quantitative differences in receptors responsible for taste synergism between MSG and GMP among three mouse strains.
INTRODIJCXION It bras been
shown
that
monosodium
strains; BALB/cCrSlc(BALB), C57BL/6CrSlc(C57BL) and C3H/HeSlc(C3H). Each mouse was anesthetized with an intraperitoneal injection of sodium pentobarbital (40-50 mg/kg), the trachea was cannulated, and the mouse was then fixed in the supine position with a headholder to allow dissection of the chorda tympani nerve. The nerve was exposed, freed from surrounding tissues and cut at the point of its entry to the bulla. For whole nerve recording, the entire nerve was placed on a silver wire electrode. An indifferent electrode was positioned nearby in the wound around the neck. Neural responses resulting from gustatory stimulation of the tongue were fed into an amplifier, integrated with a time constant of 0.1 set and displayed on a stripchart recorder. Solutions of gustatory stimulation used were 1 mM1.O M monosodium L-glutamate (MSG), 1 y M-10 mM disodium 5’-guanylate (GMP), mixtures of 0.03 M MSG and 1 PM-1GrnM GMP, mixtures of 1 mM-1.0 M MSG and 0.1 mM GMP. 0.1 M NaCl. and 0.5 M sucrose. These solutions were made up in distilldd water at about 20°C. The tongue was stimulated by flowing these test solutions at a constant flow rate (0.5 ml/set). Between two successive stimulations the tongue was rinsed with distilled water for more than 1 min to avoid possible after-effects of the preceding stimulation. To examine the property of the receptor mechanism responsible for the taste synergism between MSG and GMP, the chorda tympani responses to mixtures of MSG and GMP were compared before and after the lingual treatment of the proteolytic enzyme, 2% Pronase E (dissolved in 50 mM phosphate buffer at pH 7.0), for 10 min. For data analysis, the magnitude of the integrated chorda tympani response to each stimulus was measured 1Osec after the stimulus onset.
L-glutamate
(M:;G) elicits a unique taste, so called “umami”, to humans and produces an unusual phenomenon, the taste synergism with 5’4bonucleotides (Ikeda, 1909; Yalnaguchi, 1979). That is, when a low concentration of 5’-ribonucleotides is mixed with MSG, the taste intensity of the mixture is greater than the sum of the tastes of the components. This taste synergism between MSG and 5’-ribonucleotides has been also denlonstrated in electrophysiological studies in rats (SXO and Akaike, 1965; Sato et nl., 1970), cats (Adachi, 1964), mice (Ninomiya et al., 1989a), dogs (Kurihara, in press) and chimpanzees (Hellekant and Ninomiya, in press) examining their chroda tympani and glossopharyngeal nerve responses. However, it has been also shown that the extent of the synergism con:siderably varies among these species and among indl viduals in dogs (Kurihara, in press), and no such sym:rgistic effect is observed in the hamster chorda tympani responses (Yamamoto et al., 1988). These spel.:ies and individual differences in the receptor mechanism responsible for the taste synergism are pos,;ibly based on both genetic and environmental factors. The present study, therefore, used three inbred strams of mice, and examined the possible genetic variability of the occurrence of the taste synergism by comparing the chorda tympani responses to MSG, GMP and their mixtures among the three strains.
RESULTS
MATERIALS AND METHODS Data were obtained 8-12 weeks of age)
CBPA 101,14
from male mice of the following
Figure 1 shows sample recordings of the integrated chorda tympani responses of three mouse strains
(2&40g/b.w., three inbred 97
98
Yuzo NINOMIYAei al
h
C3H/He
-.
-
20 set 0.03M O.lmM M+G MSG GMP Fig. I. Sample recordings of the integrated chorda tympani responses of three mouse strains (BALB/c, C57BLj6 and C3H/He) to 0.03 M MSG, 0. I mM GMP and the mixture of 0.03 M MSG and 0.1 mM GMP (M + G).
(BALB, C57BL and C3H) to 0.03 M MSG, 0.1 mM GMP and the mixture of 0.03 M MSG and 0.1 mM GMP. All three mouse strains showed very good responses to the mixture, which were larger than the sum of responses to each component of the mixture, MSG and GMP, presenting the typical taste synergism between MSG and GMP. Concentrationresponse relationships of three mouse strains for MSG and mixtures of MSG and 0.1 mM GMP (M + G), and for GMP and mixtures of 0.03 M MSG and GMP are shown in Figs 2 and 3, respectively. Significant enhancement of responses to MSG by the addition of 0.1 mM GMP (over the sum of responses to each component, t-test, P -c0.05) was observed in the concentration range of MSG from 0.01 to 0.1 M in C57BL mice and from 0.01 to 1.0 M in C3H mice, but only at 0.03 M in BALB mice. Similarly, significant enhancement of 0.03 M MSG by the addition of GMP was observed in the concentration range of GMP from 0.1 to 10mM in both C57BL and C3H mice, but again only at 0.1 mM in BALB mice. Therefore, the concentration ranges of MSG and GMP, to which the synergistic responses appeared, were wider in the order of C3H > C57BL > BALB mice. Figure 4 shows per cent responses to mixtures of MSG and 0.1 mM GMP among three strains, when the sum of responses to each component was taken as unity (100%). Overall magnitudes of the synergism were apparently greater in the order of C3H > C57BL > BALB mice. The greatest synergism was observed commonly to the three strains in response to the mixture of 0.03 M MSG and 0.1 mM GMP. At this pair of concentrations, the magnitude to the mixture reached to about 260% for C3H
mice, 180% for C57BL mice and 150% for BALB mice. The lingual treatment of the proteolytic enzyme, Pronase E, suppressed responses to MSG and the mixture of MSG and GMP, and inhibited the synergism between MSG and GMP (Fig. 5). Figure 6 shows per cent responses of the chorda tympani nerve of three strains to 0.1 M NaCl, 0.5 M sucrose, 0.03 M MSG and the mixture of 0.03 M MSG and 0.1 mM GMP after the proteolytic treatment. Responses to sucrose, MSG and the mixture were significantly suppressed by pronase, but not that to NaCl, suggesting proteinaccous components of receptor mechanisms for the former substances. The magnitude of the pronase suppression of responses to both MSG and the mixture of MSG and GMP was greater in the order of C3H > C57BL> BALB mice. Per cent responses of the control to MSG and the mixture of MSG and GMP after the pronase treatment were 62.3 and 35.7% for C3H mice, 72.6 and 43.0% for C57BL mice and 81.5 and 70.1% for BALB mice, respectively. These results suggest that the receptor component responsible for the synergism between MSG and GMP is proteinaceous, and is quantitatively different among mouse strains, greater in the order of C3H > C57BL > BALB mice. DISCUSSION
Previous studies have demonstrated the existence of prominent strain differences of mice in peripheral taste nerve responses to various bitter, salty and sweet substances, such as sucrose octa-acetate (Shingai
Synergism
between
MSG
and GMP
in mice
99
BALB/c .lmM GMP)
C57BV6
9
C3WHe
Log MSG concentration (MI Fig. 2. Concentration-response relationships of three mouse strains (BALB/c, C57BL/6 and C3H/He) for MSG and mixtures of MSG and 0.1 mM GMP (M + G). Dotted line indicates the sum of responses to MSG and 0.1 mM GMP (sum). *t-test, P < 0.05; **P < 0.01; ***P < 0.001.
ancl Beidler, 1982), NaCl (Ninomiya et al., 1989b), L-P -oline and D-phenylalanine (Ninomiya et al., 1984, 1987, 1991). The present study has shown other stmin differences in taste sensitivity to an umami substance, MSG and its mixture with GMP. Three mouse strains employed in the present study were the same as those used in previous studies (Ninomiya et al., 1984, 1987, 1991) in which low sensitivities to sweet substances, such as L-proline and D-phenylalanine were reported in BALB and C3H mice as compared with that in CFBL mice. This order of mouse strains in taste sensitivities to sweet tasting amino acids was quite different from that in sensitivities to MSG and the mixture of MSG and GMP observed in the present study. This suggests different genetic basis between receptor mechanisms for these umami and sweet substances.
Several electrophysiological
studies in rats (Sato et al., 1989a) have reported that in the chorda tympani nerve almost all single fibers showing the synergism between MSG and GMP were predominantly sensitive to sucrose among the four basic taste stimuli (sucrose, NaCI, HCl and quinine), although some mouse glossopharyngeal fibers are not highly sensitive to sucrose but show the typical synergism (Ninomiya et al., 1989a). This suggests the possibility that in the chorda tympani nerve of rats and mice, neural responsiveness to sucrose and the mixture of MSG and GMP is not distinguishable, although receptor mechanisms for them are different. If this is the case, it is also possible that mouse strain differences in the magnitude of the synergism observed in the chorda tympani responses are based on those in responsiveness of sucrose-sensitive fibers to et al., 1970) and mice (Ninomiya
100
Yu20
NIN~MIYA
et at.
C57BL/6
s )_ /I
C3H/He
-3 Log GMP concentration
-2 W
Fig. 3. Concentration-response relationships of three mouse strains (BALB/c, C57BL/6 and C3H/He) for GMP and mixtures of GMP and 0.03 M MSG (M + G). Dotted line indicates the sum of responses to GMP and 0.03 M MSG (sum). *l-test, P < 0.05; **P < 0.01: ***P< 0.001.
the mixture of MSG and GNP. Unlike mice, in single fibers of the chimpanzee chorda tympani nerve showing the synergism, gymnemic acid specifically suppressed responses to sweet substances but not to the mixture of MSG and GMP (Heliekant and Ninomiya, in press). Yamamoto et al. (1988) have shown that the hamster chorda tympani and the glossopharyngeal nerves do not show the synergism between MSG and IMP or GMP and the hamster do not behaviorally disc~minate between MSG and the four basic taste stimuli. Similarly, our preliminary behavioral experiments by using a conditioned taste aversion paradigm (Ninomiya et al., in preparation) showed that behavioural discriminabilities between MSG and the four basic taste stimuli are very poor in BALB mice
showing very weak synergism in the chorda tympani responses, but are considerably higher in C57BL and C3H mice showing strong synergism. From these results, it is probable that in mice the magnitude of the synergism between MSG and GMP observed in the chorda tympani responses reflect the amount of specific receptors for the umami substances, so that behavioural discriminabilities between umami and the four basic taste stimuli are to some extent parallel with the magnitude of the synergism. However, to clarify this, since in mice the glossopharyngeal nerve is more important for behavioural disc~mination between umami and the four basic stimuli (Ninomiya et al., 1989c), it is also needed to examine taste responses of the glossopharyngeal nerve of three mouse strains.
101
Synergism between MSG and GMP in mice Percent
O.OlM
enhancement
of MSG
1O.lmMGMP
response
(sum:100
J
MSG C3H/He
0.03M
MSG SD
0.1
M
MSG
0.3
M
MSG
Fig. 4. Per cent responses to mixtures of MSG and 0.1 mM GMP in three mouse strains (BALB/c, C57BL/6 and C3H/He), when the sum of responses to each component was taken as unity (100%).
2%
Pronese(lOmin)
C57BL/6
C3H/He
-4
d Na
MSG
Irrhl
h
M+G
h hL Na
MSG
M+G
Fig. 5. Effects of the lingual treatment of 2% Pronase E for 10 min on the chorda tympani responses to 0.1 M NaCl (Na), 0.03 M MSG (MSG) and the mixture of 0.03 M MSG and 0.1 mM GMP (M + G) in three mouse strains (BALB/c, C57BL/6 and C3H/He).
zizc
102
Yuzo NINOMIYA et al. Effect
of lingual
treatment
of Pronase
-..
control
E f
2
Na
sue
MSG
M+G
Fig. 6. Per cent responses to 0.1 M NaCl (Na), 0.5 M sucrose (Sue), 0.03 M MSG (MSG and the mixture of 0.03 M MSG and 0.1 mM GMP (M + G) after the lingual treatment of 2% Pronase E for 10 min in three mouse strains (BALB/c, C57BL/6 and C3H/He). Control = 100%. *r-test, P < 0.05; **P < 0.01; ***p < 0.001. Acknowledgements-This study was supported in part by Grants-in-Aid for Scientific Research (No. 63480418) from the Ministry of Education, Science and Culture of Japan, REFERENCES
Adachi A. (1964) Neurophysiological study on taste effectiveness of seasoning. J. Physiol. Sot. Jpn 26, 347-355. Hellekant G. and Ninomiya Y. On the taste of umami in chimpanzee. Physiol. Behau. (In press). Ikeda K. (1909) On a new seasoning. J. Tokyo Chem. Sot. 30, 82G-836 (in Japanese). Kurihara K. Large synergism between MSG and nucleotides and differentiation between umami and salt components in canine taste nerve responses. Physiol. Behau. (In press). Ninomiya Y., Mizukoshi T., Higashi T.. Katsukawa H. and Funakoshi M. (1984) Gustatory neural responses in three different strains of mice. Brain Res. 302,305S3 14.
Ninomiya Y., Higashi T., Mizukoshi T. and Funakoshi M. (1987) Genetics of the ability to perceive sweetness of o-phenylalanine in mice. Ann. NY Acad. Sci. 510, 527-529. Ninomiya Y. and Funakoshi M. (1989a) Peripheral neural basis for behavioural discrimination between glutamate and the four basic taste substances in mice. Comp. Biochem. Physiol. 92, 371-376. Ninomiya Y., Sako N. and Funakoshi M. (1989b) Strain differences in amiloride inhibition of NaCl responses in mice, Mus musealas. J. camp. Physiol. 166, l-5. Ninomiya Y. and Funakoshi M. (1989~) Behavioural discrimination between glutamate and the four basic taste substances in mice. Comp. Biochem. Physiol. 92, 365-370. Ninomiya Y., Sako N., Katsukawa H. and Funakoshi M. (1991) Taste receptor mechanisms influenced by a gene on chromosome 4 in mice. In Chemical Senses: Vol. 3: Genetics of Perception and Communication (Edited by Wysocki C. and Kare M. R.), pp. 2677278. Marcel Dekker, New York. Sato M. and Akaike N. (1965) 5’-ribonucleotides as gustatory stimuli in rats. Electrophysiological studies. Jpn J. Physiol. 15, 53-70. Sato M., Yamashita S. and Ogawa H. (1970) Potentiation of gustatory response to monosodium glutamate in rat chorda tympani fibers by addition of 5’-ribonucleotides. Jpn J. Physiol. 20, 444464. Shingai T. and Beidler L. M. (1985) Response characteristics of three taste nerves in mice. Bruin Res. 335, 2455249. Yamaguchi S. (1979) The umami taste. In Food Taste Chemistry (Edited by Boudreau J. C.), pp. 33-5 I. American Chemical Society, Washington, DC. Yamamoto T., Matsuo R., Kiyomitsu Y. and Kitamura R. (1988) Taste effects of umami substances in hamsters as studied by electrophysiological and conditioned taste aversion techniques. Brain Res. 451, 147-162.
Camp. Eiochem. Physiol. Vol. IOIA, No. I, pp. 97-102, 1992 Primed in Great Britain
0
1991 Pergamon
Press plc
TASTE SYNERGISM BETWEEN MONOSODIUM GLUTAMATE AND 5’-RIBONUCLEOTIDE IN MICE Yuzo
NINOMIYA, SHUJI KURENUMA,* TAKAYUKI NOMURA, HAJIMU UEBAYASHI and HIROSADA KAWAMURA
Department
of Oral Physiology and *Oral Surgery, Asahi University School of Dentistry, Hozumi, Motosu, Gifu 501-02, Japan. Telephone: 05832-6-613 1 (Received
17 April
1991)
Abstract-l. Strain differences of mice were found in the taste synergism between monosodium L-glutamate (MSG) and disodium 5’-guanylate (GMP). 2. Magnitudes of chorda tympani responses to the mixture of MSG and GMP over the sum of responses to each component were greater in the order of C3H/HeSlc(C3H) > C57BL/6CrSlc(C57BL) > BALB/cCrSlc(BALB) mice. The greatest synergism was observed in response to the mixture of 0.03 M MSG and 0.1 mM GMP, to which responses were about 2.6, 1.8 and 1.4 times greater than the sum of each component in C3H, C57BL and BALB mice, respectively. 3. Magnitudes of inhibition of MSG and mixture responses by the lingual treatment of proteolytic enzyme, Pronase E, were greater in the same order of C3H > C57BL > BALB mice as that observed in magnitudes of the synergism. These results suggest that there exists quantitative differences in receptors responsible for taste synergism between MSG and GMP among three mouse strains.
INTRODIJCXION It bras been
shown
that
monosodium
strains; BALB/cCrSlc(BALB), C57BL/6CrSlc(C57BL) and C3H/HeSlc(C3H). Each mouse was anesthetized with an intraperitoneal injection of sodium pentobarbital (40-50 mg/kg), the trachea was cannulated, and the mouse was then fixed in the supine position with a headholder to allow dissection of the chorda tympani nerve. The nerve was exposed, freed from surrounding tissues and cut at the point of its entry to the bulla. For whole nerve recording, the entire nerve was placed on a silver wire electrode. An indifferent electrode was positioned nearby in the wound around the neck. Neural responses resulting from gustatory stimulation of the tongue were fed into an amplifier, integrated with a time constant of 0.1 set and displayed on a stripchart recorder. Solutions of gustatory stimulation used were 1 mM1.O M monosodium L-glutamate (MSG), 1 y M-10 mM disodium 5’-guanylate (GMP), mixtures of 0.03 M MSG and 1 PM-1GrnM GMP, mixtures of 1 mM-1.0 M MSG and 0.1 mM GMP. 0.1 M NaCl. and 0.5 M sucrose. These solutions were made up in distilldd water at about 20°C. The tongue was stimulated by flowing these test solutions at a constant flow rate (0.5 ml/set). Between two successive stimulations the tongue was rinsed with distilled water for more than 1 min to avoid possible after-effects of the preceding stimulation. To examine the property of the receptor mechanism responsible for the taste synergism between MSG and GMP, the chorda tympani responses to mixtures of MSG and GMP were compared before and after the lingual treatment of the proteolytic enzyme, 2% Pronase E (dissolved in 50 mM phosphate buffer at pH 7.0), for 10 min. For data analysis, the magnitude of the integrated chorda tympani response to each stimulus was measured 1Osec after the stimulus onset.
L-glutamate
(M:;G) elicits a unique taste, so called “umami”, to humans and produces an unusual phenomenon, the taste synergism with 5’4bonucleotides (Ikeda, 1909; Yalnaguchi, 1979). That is, when a low concentration of 5’-ribonucleotides is mixed with MSG, the taste intensity of the mixture is greater than the sum of the tastes of the components. This taste synergism between MSG and 5’-ribonucleotides has been also denlonstrated in electrophysiological studies in rats (SXO and Akaike, 1965; Sato et nl., 1970), cats (Adachi, 1964), mice (Ninomiya et al., 1989a), dogs (Kurihara, in press) and chimpanzees (Hellekant and Ninomiya, in press) examining their chroda tympani and glossopharyngeal nerve responses. However, it has been also shown that the extent of the synergism con:siderably varies among these species and among indl viduals in dogs (Kurihara, in press), and no such sym:rgistic effect is observed in the hamster chorda tympani responses (Yamamoto et al., 1988). These spel.:ies and individual differences in the receptor mechanism responsible for the taste synergism are pos,;ibly based on both genetic and environmental factors. The present study, therefore, used three inbred strams of mice, and examined the possible genetic variability of the occurrence of the taste synergism by comparing the chorda tympani responses to MSG, GMP and their mixtures among the three strains.
RESULTS
MATERIALS AND METHODS Data were obtained 8-12 weeks of age)
CBPA 101,14
from male mice of the following
Figure 1 shows sample recordings of the integrated chorda tympani responses of three mouse strains
(2&40g/b.w., three inbred 97
98
Yuzo NINOMIYAei al
h
C3H/He
-.
-
20 set 0.03M O.lmM M+G MSG GMP Fig. I. Sample recordings of the integrated chorda tympani responses of three mouse strains (BALB/c, C57BLj6 and C3H/He) to 0.03 M MSG, 0. I mM GMP and the mixture of 0.03 M MSG and 0.1 mM GMP (M + G).
(BALB, C57BL and C3H) to 0.03 M MSG, 0.1 mM GMP and the mixture of 0.03 M MSG and 0.1 mM GMP. All three mouse strains showed very good responses to the mixture, which were larger than the sum of responses to each component of the mixture, MSG and GMP, presenting the typical taste synergism between MSG and GMP. Concentrationresponse relationships of three mouse strains for MSG and mixtures of MSG and 0.1 mM GMP (M + G), and for GMP and mixtures of 0.03 M MSG and GMP are shown in Figs 2 and 3, respectively. Significant enhancement of responses to MSG by the addition of 0.1 mM GMP (over the sum of responses to each component, t-test, P -c0.05) was observed in the concentration range of MSG from 0.01 to 0.1 M in C57BL mice and from 0.01 to 1.0 M in C3H mice, but only at 0.03 M in BALB mice. Similarly, significant enhancement of 0.03 M MSG by the addition of GMP was observed in the concentration range of GMP from 0.1 to 10mM in both C57BL and C3H mice, but again only at 0.1 mM in BALB mice. Therefore, the concentration ranges of MSG and GMP, to which the synergistic responses appeared, were wider in the order of C3H > C57BL > BALB mice. Figure 4 shows per cent responses to mixtures of MSG and 0.1 mM GMP among three strains, when the sum of responses to each component was taken as unity (100%). Overall magnitudes of the synergism were apparently greater in the order of C3H > C57BL > BALB mice. The greatest synergism was observed commonly to the three strains in response to the mixture of 0.03 M MSG and 0.1 mM GMP. At this pair of concentrations, the magnitude to the mixture reached to about 260% for C3H
mice, 180% for C57BL mice and 150% for BALB mice. The lingual treatment of the proteolytic enzyme, Pronase E, suppressed responses to MSG and the mixture of MSG and GMP, and inhibited the synergism between MSG and GMP (Fig. 5). Figure 6 shows per cent responses of the chorda tympani nerve of three strains to 0.1 M NaCl, 0.5 M sucrose, 0.03 M MSG and the mixture of 0.03 M MSG and 0.1 mM GMP after the proteolytic treatment. Responses to sucrose, MSG and the mixture were significantly suppressed by pronase, but not that to NaCl, suggesting proteinaccous components of receptor mechanisms for the former substances. The magnitude of the pronase suppression of responses to both MSG and the mixture of MSG and GMP was greater in the order of C3H > C57BL> BALB mice. Per cent responses of the control to MSG and the mixture of MSG and GMP after the pronase treatment were 62.3 and 35.7% for C3H mice, 72.6 and 43.0% for C57BL mice and 81.5 and 70.1% for BALB mice, respectively. These results suggest that the receptor component responsible for the synergism between MSG and GMP is proteinaceous, and is quantitatively different among mouse strains, greater in the order of C3H > C57BL > BALB mice. DISCUSSION
Previous studies have demonstrated the existence of prominent strain differences of mice in peripheral taste nerve responses to various bitter, salty and sweet substances, such as sucrose octa-acetate (Shingai
Synergism
between
MSG
and GMP
in mice
99
BALB/c .lmM GMP)
C57BV6
9
C3WHe
Log MSG concentration (MI Fig. 2. Concentration-response relationships of three mouse strains (BALB/c, C57BL/6 and C3H/He) for MSG and mixtures of MSG and 0.1 mM GMP (M + G). Dotted line indicates the sum of responses to MSG and 0.1 mM GMP (sum). *t-test, P < 0.05; **P < 0.01; ***P < 0.001.
ancl Beidler, 1982), NaCl (Ninomiya et al., 1989b), L-P -oline and D-phenylalanine (Ninomiya et al., 1984, 1987, 1991). The present study has shown other stmin differences in taste sensitivity to an umami substance, MSG and its mixture with GMP. Three mouse strains employed in the present study were the same as those used in previous studies (Ninomiya et al., 1984, 1987, 1991) in which low sensitivities to sweet substances, such as L-proline and D-phenylalanine were reported in BALB and C3H mice as compared with that in CFBL mice. This order of mouse strains in taste sensitivities to sweet tasting amino acids was quite different from that in sensitivities to MSG and the mixture of MSG and GMP observed in the present study. This suggests different genetic basis between receptor mechanisms for these umami and sweet substances.
Several electrophysiological
studies in rats (Sato et al., 1989a) have reported that in the chorda tympani nerve almost all single fibers showing the synergism between MSG and GMP were predominantly sensitive to sucrose among the four basic taste stimuli (sucrose, NaCI, HCl and quinine), although some mouse glossopharyngeal fibers are not highly sensitive to sucrose but show the typical synergism (Ninomiya et al., 1989a). This suggests the possibility that in the chorda tympani nerve of rats and mice, neural responsiveness to sucrose and the mixture of MSG and GMP is not distinguishable, although receptor mechanisms for them are different. If this is the case, it is also possible that mouse strain differences in the magnitude of the synergism observed in the chorda tympani responses are based on those in responsiveness of sucrose-sensitive fibers to et al., 1970) and mice (Ninomiya
100
Yu20
NIN~MIYA
et at.
C57BL/6
s )_ /I
C3H/He
-3 Log GMP concentration
-2 W
Fig. 3. Concentration-response relationships of three mouse strains (BALB/c, C57BL/6 and C3H/He) for GMP and mixtures of GMP and 0.03 M MSG (M + G). Dotted line indicates the sum of responses to GMP and 0.03 M MSG (sum). *l-test, P < 0.05; **P < 0.01: ***P< 0.001.
the mixture of MSG and GNP. Unlike mice, in single fibers of the chimpanzee chorda tympani nerve showing the synergism, gymnemic acid specifically suppressed responses to sweet substances but not to the mixture of MSG and GMP (Heliekant and Ninomiya, in press). Yamamoto et al. (1988) have shown that the hamster chorda tympani and the glossopharyngeal nerves do not show the synergism between MSG and IMP or GMP and the hamster do not behaviorally disc~minate between MSG and the four basic taste stimuli. Similarly, our preliminary behavioral experiments by using a conditioned taste aversion paradigm (Ninomiya et al., in preparation) showed that behavioural discriminabilities between MSG and the four basic taste stimuli are very poor in BALB mice
showing very weak synergism in the chorda tympani responses, but are considerably higher in C57BL and C3H mice showing strong synergism. From these results, it is probable that in mice the magnitude of the synergism between MSG and GMP observed in the chorda tympani responses reflect the amount of specific receptors for the umami substances, so that behavioural discriminabilities between umami and the four basic taste stimuli are to some extent parallel with the magnitude of the synergism. However, to clarify this, since in mice the glossopharyngeal nerve is more important for behavioural disc~mination between umami and the four basic stimuli (Ninomiya et al., 1989c), it is also needed to examine taste responses of the glossopharyngeal nerve of three mouse strains.
101
Synergism between MSG and GMP in mice Percent
O.OlM
enhancement
of MSG
1O.lmMGMP
response
(sum:100
J
MSG C3H/He
0.03M
MSG SD
0.1
M
MSG
0.3
M
MSG
Fig. 4. Per cent responses to mixtures of MSG and 0.1 mM GMP in three mouse strains (BALB/c, C57BL/6 and C3H/He), when the sum of responses to each component was taken as unity (100%).
2%
Pronese(lOmin)
C57BL/6
C3H/He
-4
d Na
MSG
Irrhl
h
M+G
h hL Na
MSG
M+G
Fig. 5. Effects of the lingual treatment of 2% Pronase E for 10 min on the chorda tympani responses to 0.1 M NaCl (Na), 0.03 M MSG (MSG) and the mixture of 0.03 M MSG and 0.1 mM GMP (M + G) in three mouse strains (BALB/c, C57BL/6 and C3H/He).
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Yuzo NINOMIYA et al. Effect
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Fig. 6. Per cent responses to 0.1 M NaCl (Na), 0.5 M sucrose (Sue), 0.03 M MSG (MSG and the mixture of 0.03 M MSG and 0.1 mM GMP (M + G) after the lingual treatment of 2% Pronase E for 10 min in three mouse strains (BALB/c, C57BL/6 and C3H/He). Control = 100%. *r-test, P < 0.05; **P < 0.01; ***p < 0.001. Acknowledgements-This study was supported in part by Grants-in-Aid for Scientific Research (No. 63480418) from the Ministry of Education, Science and Culture of Japan, REFERENCES
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