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Chirality in insect chemoreceptors
Pargamoa Pras
Life Saieaaee Vol. 14, pp " 2045-2049 Printed is II.S .A .
CH7RALITY IN INSECT CHEMORECEPTORB Yaaoov Lensky and Murray S . Blam D~partment of Eatomolo®, Uaiverelty of (ieorgie, Atheae, Cleorgls 30808 (Reo~ived in final form 22 April 1974) Sammsry Canditiaaed honeybee workers osa disoriminabe between ensatiomerio paire which are both oangruoue and inoooglvous odorante for human beings . These reealta are dleoassed in terms ad appropriately ohlral ohemoreoeptore, which would enable honey_ bee workers bo distinguish floral odor eoaroee that differ aniy in enaatiomerio identity . A variety of pheromones syatheeised by insects contain centers of ohirality (1, 8, 3, 4, 6, 8) . in view of the enantiomerio etereospecifiaity that oharacterlsee en$ymee sad proteins la go~ral, !t is possible that meat' pheromones are synthesised with a ohiral exactitude.
Furthermore, the ohemoreoeptore that dlaorlminabe these isomers
are likely to possess as appropriate ahirality. 8lgniâoa~y, the presence of ohemoreceptors with a well-devela~ped eterio enantiomorphiem would enable insects to attain maximum ~electivity to the mnltitade of ohiral natural products which lhmction as pleat attraateats, phagostlmalaats and chemical messengers .
Indeed, the eeleotivity of ia-
sect ohemoreoeptors for single enantiomere may result is a greater sensitivity to chiral pheromones thaw can be determined by evaluating their response to racemio min tares ('n . This report esaminee the ability of honeybee workers to discriminate enaatiomerio pairs, and in partloa]ar : 1. Enaatiomere which possess inaongruoue odors for humane :
(R)- and
(S)-oarvoae (8, 9,10) . 8 . An anantiomer and a raoemio mixt
(R)= and
204 6
Chiralitp in Chemorsc~ptors
Vol . 14, No. 10
The results of this study demaastrabe that honeybees aaa readily discrimhiate between both pairs of enaatlomere and thus probably are aapsble of responding to a variety of chiral environmental "triggers" .
Materials and Methods The ai)- and (S)-enaatiomere of aarvone were synthesized by Dr. L. Friedmsn, Case-Western Reserve University, Cleveland and the (R)- and (S)-enaatlomere of 2-oatanol were provided by Dr. J. (3. Trayaham, Louisiana State University. The de
gree of purity of these aompaunds was 99~ and 98~, respectively. The fimatioa of insect olfactory organs aaa be studied through observations of behavioral responses to a given stimulus, sad in the honeybee the proboscis reflex ie used as a means of measuring the degree of aanditianb~g to a speoi$c odor (11) . This behavioral response to odor stimulation is more speoiüo than that of the receptor aelle which trigger the response (12). 1n our study, honeybee worksre were trained is the laboratory to indicate their assoaiaticn of a specilYc odor with the availsbüity of food, by extending the proboscis . Their ability to disoriminats between the known and unknown aompounde was thus made manifest. üalisa honeybee A is mellifera L. var . 1 iguetioa Spin. ) field workers were aasssthetized with C02 and were transferred to plastic cylinders . Only their heads were exposed (1S). The base were aanditioned to the trabiing odor and subseque~y rewarded with a 90~ aqueous honey solution. Following the establishment of a
"memory" to the training odor, the bees eabended their proboscis in the presence of the odor substance without being rewarded with honey solatian . A full description of the method will be published elsewhere . The besting procedure bicluded the following four oaoseoutive exposures (each
of 10 sea . ) in each repetition: (a) air only: (b) air and training substance ; (o) air calyp (d) air and the candidate substance . A trained bee, which was able to discriminate between the two enantiomers, would extend its proboscis only to the training compound (easatiomer) but not when exposed to fire other candidate isomer . The "memory" for the training stimulus persisted for several hours during which time about 16 to 20 oanseoutive tests were carried out. Results and Disauseion Bees which were trained to (S)-aarvone diearlmiasted almost perfectly between the two emntiomere. Ia a total of 44 tests, 41 positive results to etimalatian by
vol. 14, No . 10
Chiralitp ia Chrooreapàors
soa~
(~-oarvone were recorded, but and 8 to (Rhoarvane . Heee trained to (8~-oarvaoe
responded to it positivelyy m a total of ao tests ~ad distinguished it lSrom (:~-oarvane m
ao additicoal tests. When bees were framed to ~)-oarvcoe they reaobed positively to stimnlatian b9 a raoemio mizbue (ia ; v/v) of (ßF and (!~-oarvone . However, the daratian ad probosota eabension to the raoemio mixture was shorter m several fieteaoee than to ~)-oarvane alone . The resuüs demonstrate that honeybee workers are able to distmgnish between
the odors of the (~~matiomer and the ~)-enantiomer. To the human nose these
ocmpamds smell ad epearmmt and caraway, respectively. No mterferenae with the perception of the adore occurred when the workers were eaposad to the raoemio miztare. Whereas the diäere~e between the odors of the (R)- and (:;)-oarvone ensatiomere to humane ie hadispukable (S, 9,10), the ~)- ~ad ~-eoaatiomers ad 2-ootaaol oam~ot be distmguiehed by honnsae (8). m contrast, bee worloere framed to ~)-S-
ootanol readily dietmguiehed between the two eneatiomere, by extending the proboscis to the (R)-, but not to the (8}-easatiomer, In the 81 tests performed.
û thus appears that the honeybee worker oaa dieo r+w,i,+~" at least some enaatio-
mere with greater eouity thaw hnmsa beings . Cooeidertng the widespread ooourrenoe od optloal isomers ae volatile products of plants (14), it should oak be eurprlemg that honeybees have evolved ohemoreoeptore capable os acting spedtloaliy to these o6lral
oompoande, maa
laotto acid, e~ it is signitioaat that the ~)-lactic acid is the mom component m mammalian sweat (16) .
The distinction between ensatiomere of oarvones sad &~ootaaols hoe been demm-
etrated in oar ezpertments oeiug uadilabed compounds. û should be emphasised that
the behavioral reaoücn apt honeybees (proboscis extension) that we observed was eeeentially all or none.
m all probability, the honeybee workers utillrs their ability to discriminate enaatiomere which are employed m mtraspeoiflo oommaatoation . For example, one of the gases bee pheramooee, 8-hydraaq- trace-8-deoenio acid, which !~motions to
etabdlise worker swarms Cl), oanteine a obis! center, sad the great sensitivity of workers to this compamd may reIIect their selective response to a naturally occurring enaatiamer.
Oar data do not support the vibrattooal theory ad odor (18) which correlates the odor qualities of compounds with epeoiflo vibrations! beads in the far-h~frared region.
204 8
Chirality in Chemoreceptore
Vol . 14, No . 10
Differences in the vibratlaasl and internal energies of enantlomers would be identical, resulting in oangruent far-Infrared spectra of the optical isomers .
The elbrational
theory ie also hsoompatible with the demonstration that ineeota, including the honeybee, cannot distinguish deuterated adorant molecules from their non-deuterated oountsxL parts (17,18,19), notwithetaadlng the shifts in the far- in~~ absorption ma~dma that reealt when deuterium ie hstroduoed into these compounds . The demonstration that a difference in chirality of optical isomers released a clear-cut behavioral response to honeybee workers (proboscis extension), notwithstanding the fact that none of the enantiomers examined by us was a natural product of the honeybee (pheromone or alarm substance), would suggest the eudeteaoe of appropriately chiral ahemoreoeptora in insects .
indeed, moat biochemical reactions
are etereoepecifio and although the mechanism of recognition of odor molecules by epeeific receptors is unlmowa, there ie experimental eeidenae suggesting that specific receptor proteins are involved in this discrimination (20) . Signiflcant),y, our remits demonstrate that honeybee workers are emhzent,)~y capable of storing the information encoded in enantiomers in the central nervous system as a highlvy adaptive device for responding to chemical signals identified with specific food sources .
Thus, the presence of ahiral ahemoreceptors for discriminating optical
isomers, coupled with the ability to "memorize" the information encoded in these specific signals, has probably permitted these insects to aucceseitiilly exploit the multitude ad ohirally dependent cues emanating from food plants . Acknowledgments We wish to express our gratitude to Professor C . Teso for providing experimentalfaoilities and Professor A . Dletz for providing honeybees . References 1.
C . G . BUTLER, R . K . CALLOW and J. R . CHAPMAN, Nature 201, 755 (1984) .
2.
D. J . MoGURK, J . FROST, E . J . EISENBRAUN, K . VICK, W . A . DREW sad J. YOUNG, J. Insect
sioL 12, 1956 (1988) .
5.
R. M . CREWE and M. S . BLUM, J. Insect P
eiol. 18, 141 (1970) .
4.
C . W . CAVILL and W . HIDiTERBERGER, Proo . Wit. Caogr . Ent . 11th Vienna 5, 65 (1882) .
6.
Ii. M . FALES, M. S . BLUM, R . M . CREWE and J . M . BRAND, J . Insect elol. 1 1077 (1972) .
204 9
Chiralitp ia Ch~orecspton
Vol . 14, No . 10
8.
R. M . SILVERSTEIN, J . O. RODIId sad D . L . WOOD, 8oienoe 164, 609 (1988) .
7.
J . C . MOSER, R . G . BROWNLSE aad R . M. SILVERSTEIId, J. Ineeot
iol.
14, 629 (1988) . 8.
L. FRIEDMAN aad J . G . MII~LER, Scienoe 172, 1044 (1971) .
9.
G. F . RUSSELL aad J . L HILLS, Soienoe 172. 1043 (1971) .
10 .
T. J. LEITEREG, D. G. GUAUDAGNI, JEAN HARRIB, T. R. MON aad R . TERANIBHI, Nature 230, 466 (1971) .
il.
K. TAKEDA, J . Ineeot
siol . 8, 188 (1981) .
12.
B. STLTRCKOW, in : Advanaee in Chemoreoeptian (Eds : J. W. JOBNSTON, Jr. , D. G . MOULTON, and A . TURK) Vol . York, 1970) .
1
107 (Appletan-Century-Cra~lts, New
iol. 13. E . VARESCHI, Z . ve . 7b, 143 (1971) . 14.
W . KARRER, Kaaetitutim :md Vorkommen der Organisohen P1lsazenetodfs , (Birkhaaeer Verlag, Berlin, 1968) .
i6.
F. ACREE, Jr., R. B . TURNER, H. K . GOUCK, M . BEROZA aad N. BMITH, Soienoe 181, 1348 (1988) .
18 .
R . H. WRIGHT, The 8oienoe of 8mell, (Baeia Booka, N . Y. , 1984) .
17.
R . J. BAKER, R. L . BERDEL and G . D. WALKER, E~erientla 29 . 418 (1973) .
18.
R. E . DOOLITTLE, M. BEROZA, I . KEIBER and E . L. SCHNEIDER, J . fiseot eiol. 14, 1897 (1973) .
19.
M . 8 . BLUM, R. E . DOOLTITLE and M . BEROZA, J, mseot P 2361 (1971) .
20.
L. M. RIDDII+'ORD, J. >hseot
eiol . 18, 863 (1970) .
siol . 17,
Life Saieaaee Vol. 14, pp " 2045-2049 Printed is II.S .A .
CH7RALITY IN INSECT CHEMORECEPTORB Yaaoov Lensky and Murray S . Blam D~partment of Eatomolo®, Uaiverelty of (ieorgie, Atheae, Cleorgls 30808 (Reo~ived in final form 22 April 1974) Sammsry Canditiaaed honeybee workers osa disoriminabe between ensatiomerio paire which are both oangruoue and inoooglvous odorante for human beings . These reealta are dleoassed in terms ad appropriately ohlral ohemoreoeptore, which would enable honey_ bee workers bo distinguish floral odor eoaroee that differ aniy in enaatiomerio identity . A variety of pheromones syatheeised by insects contain centers of ohirality (1, 8, 3, 4, 6, 8) . in view of the enantiomerio etereospecifiaity that oharacterlsee en$ymee sad proteins la go~ral, !t is possible that meat' pheromones are synthesised with a ohiral exactitude.
Furthermore, the ohemoreoeptore that dlaorlminabe these isomers
are likely to possess as appropriate ahirality. 8lgniâoa~y, the presence of ohemoreceptors with a well-devela~ped eterio enantiomorphiem would enable insects to attain maximum ~electivity to the mnltitade of ohiral natural products which lhmction as pleat attraateats, phagostlmalaats and chemical messengers .
Indeed, the eeleotivity of ia-
sect ohemoreoeptors for single enantiomere may result is a greater sensitivity to chiral pheromones thaw can be determined by evaluating their response to racemio min tares ('n . This report esaminee the ability of honeybee workers to discriminate enaatiomerio pairs, and in partloa]ar : 1. Enaatiomere which possess inaongruoue odors for humane :
(R)- and
(S)-oarvoae (8, 9,10) . 8 . An anantiomer and a raoemio mixt
(R)= and
204 6
Chiralitp in Chemorsc~ptors
Vol . 14, No. 10
The results of this study demaastrabe that honeybees aaa readily discrimhiate between both pairs of enaatlomere and thus probably are aapsble of responding to a variety of chiral environmental "triggers" .
Materials and Methods The ai)- and (S)-enaatiomere of aarvone were synthesized by Dr. L. Friedmsn, Case-Western Reserve University, Cleveland and the (R)- and (S)-enaatlomere of 2-oatanol were provided by Dr. J. (3. Trayaham, Louisiana State University. The de
gree of purity of these aompaunds was 99~ and 98~, respectively. The fimatioa of insect olfactory organs aaa be studied through observations of behavioral responses to a given stimulus, sad in the honeybee the proboscis reflex ie used as a means of measuring the degree of aanditianb~g to a speoi$c odor (11) . This behavioral response to odor stimulation is more speoiüo than that of the receptor aelle which trigger the response (12). 1n our study, honeybee worksre were trained is the laboratory to indicate their assoaiaticn of a specilYc odor with the availsbüity of food, by extending the proboscis . Their ability to disoriminats between the known and unknown aompounde was thus made manifest. üalisa honeybee A is mellifera L. var . 1 iguetioa Spin. ) field workers were aasssthetized with C02 and were transferred to plastic cylinders . Only their heads were exposed (1S). The base were aanditioned to the trabiing odor and subseque~y rewarded with a 90~ aqueous honey solution. Following the establishment of a
"memory" to the training odor, the bees eabended their proboscis in the presence of the odor substance without being rewarded with honey solatian . A full description of the method will be published elsewhere . The besting procedure bicluded the following four oaoseoutive exposures (each
of 10 sea . ) in each repetition: (a) air only: (b) air and training substance ; (o) air calyp (d) air and the candidate substance . A trained bee, which was able to discriminate between the two enantiomers, would extend its proboscis only to the training compound (easatiomer) but not when exposed to fire other candidate isomer . The "memory" for the training stimulus persisted for several hours during which time about 16 to 20 oanseoutive tests were carried out. Results and Disauseion Bees which were trained to (S)-aarvone diearlmiasted almost perfectly between the two emntiomere. Ia a total of 44 tests, 41 positive results to etimalatian by
vol. 14, No . 10
Chiralitp ia Chrooreapàors
soa~
(~-oarvone were recorded, but and 8 to (Rhoarvane . Heee trained to (8~-oarvaoe
responded to it positivelyy m a total of ao tests ~ad distinguished it lSrom (:~-oarvane m
ao additicoal tests. When bees were framed to ~)-oarvcoe they reaobed positively to stimnlatian b9 a raoemio mizbue (ia ; v/v) of (ßF and (!~-oarvone . However, the daratian ad probosota eabension to the raoemio mixture was shorter m several fieteaoee than to ~)-oarvane alone . The resuüs demonstrate that honeybee workers are able to distmgnish between
the odors of the (~~matiomer and the ~)-enantiomer. To the human nose these
ocmpamds smell ad epearmmt and caraway, respectively. No mterferenae with the perception of the adore occurred when the workers were eaposad to the raoemio miztare. Whereas the diäere~e between the odors of the (R)- and (:;)-oarvone ensatiomere to humane ie hadispukable (S, 9,10), the ~)- ~ad ~-eoaatiomers ad 2-ootaaol oam~ot be distmguiehed by honnsae (8). m contrast, bee worloere framed to ~)-S-
ootanol readily dietmguiehed between the two eneatiomere, by extending the proboscis to the (R)-, but not to the (8}-easatiomer, In the 81 tests performed.
û thus appears that the honeybee worker oaa dieo r+w,i,+~" at least some enaatio-
mere with greater eouity thaw hnmsa beings . Cooeidertng the widespread ooourrenoe od optloal isomers ae volatile products of plants (14), it should oak be eurprlemg that honeybees have evolved ohemoreoeptore capable os acting spedtloaliy to these o6lral
oompoande, maa
laotto acid, e~ it is signitioaat that the ~)-lactic acid is the mom component m mammalian sweat (16) .
The distinction between ensatiomere of oarvones sad &~ootaaols hoe been demm-
etrated in oar ezpertments oeiug uadilabed compounds. û should be emphasised that
the behavioral reaoücn apt honeybees (proboscis extension) that we observed was eeeentially all or none.
m all probability, the honeybee workers utillrs their ability to discriminate enaatiomere which are employed m mtraspeoiflo oommaatoation . For example, one of the gases bee pheramooee, 8-hydraaq- trace-8-deoenio acid, which !~motions to
etabdlise worker swarms Cl), oanteine a obis! center, sad the great sensitivity of workers to this compamd may reIIect their selective response to a naturally occurring enaatiamer.
Oar data do not support the vibrattooal theory ad odor (18) which correlates the odor qualities of compounds with epeoiflo vibrations! beads in the far-h~frared region.
204 8
Chirality in Chemoreceptore
Vol . 14, No . 10
Differences in the vibratlaasl and internal energies of enantlomers would be identical, resulting in oangruent far-Infrared spectra of the optical isomers .
The elbrational
theory ie also hsoompatible with the demonstration that ineeota, including the honeybee, cannot distinguish deuterated adorant molecules from their non-deuterated oountsxL parts (17,18,19), notwithetaadlng the shifts in the far- in~~ absorption ma~dma that reealt when deuterium ie hstroduoed into these compounds . The demonstration that a difference in chirality of optical isomers released a clear-cut behavioral response to honeybee workers (proboscis extension), notwithstanding the fact that none of the enantiomers examined by us was a natural product of the honeybee (pheromone or alarm substance), would suggest the eudeteaoe of appropriately chiral ahemoreoeptora in insects .
indeed, moat biochemical reactions
are etereoepecifio and although the mechanism of recognition of odor molecules by epeeific receptors is unlmowa, there ie experimental eeidenae suggesting that specific receptor proteins are involved in this discrimination (20) . Signiflcant),y, our remits demonstrate that honeybee workers are emhzent,)~y capable of storing the information encoded in enantiomers in the central nervous system as a highlvy adaptive device for responding to chemical signals identified with specific food sources .
Thus, the presence of ahiral ahemoreceptors for discriminating optical
isomers, coupled with the ability to "memorize" the information encoded in these specific signals, has probably permitted these insects to aucceseitiilly exploit the multitude ad ohirally dependent cues emanating from food plants . Acknowledgments We wish to express our gratitude to Professor C . Teso for providing experimentalfaoilities and Professor A . Dletz for providing honeybees . References 1.
C . G . BUTLER, R . K . CALLOW and J. R . CHAPMAN, Nature 201, 755 (1984) .
2.
D. J . MoGURK, J . FROST, E . J . EISENBRAUN, K . VICK, W . A . DREW sad J. YOUNG, J. Insect
sioL 12, 1956 (1988) .
5.
R. M . CREWE and M. S . BLUM, J. Insect P
eiol. 18, 141 (1970) .
4.
C . W . CAVILL and W . HIDiTERBERGER, Proo . Wit. Caogr . Ent . 11th Vienna 5, 65 (1882) .
6.
Ii. M . FALES, M. S . BLUM, R . M . CREWE and J . M . BRAND, J . Insect elol. 1 1077 (1972) .
204 9
Chiralitp ia Ch~orecspton
Vol . 14, No . 10
8.
R. M . SILVERSTEIN, J . O. RODIId sad D . L . WOOD, 8oienoe 164, 609 (1988) .
7.
J . C . MOSER, R . G . BROWNLSE aad R . M. SILVERSTEIId, J. Ineeot
iol.
14, 629 (1988) . 8.
L. FRIEDMAN aad J . G . MII~LER, Scienoe 172, 1044 (1971) .
9.
G. F . RUSSELL aad J . L HILLS, Soienoe 172. 1043 (1971) .
10 .
T. J. LEITEREG, D. G. GUAUDAGNI, JEAN HARRIB, T. R. MON aad R . TERANIBHI, Nature 230, 466 (1971) .
il.
K. TAKEDA, J . Ineeot
siol . 8, 188 (1981) .
12.
B. STLTRCKOW, in : Advanaee in Chemoreoeptian (Eds : J. W. JOBNSTON, Jr. , D. G . MOULTON, and A . TURK) Vol . York, 1970) .
1
107 (Appletan-Century-Cra~lts, New
iol. 13. E . VARESCHI, Z . ve . 7b, 143 (1971) . 14.
W . KARRER, Kaaetitutim :md Vorkommen der Organisohen P1lsazenetodfs , (Birkhaaeer Verlag, Berlin, 1968) .
i6.
F. ACREE, Jr., R. B . TURNER, H. K . GOUCK, M . BEROZA aad N. BMITH, Soienoe 181, 1348 (1988) .
18 .
R . H. WRIGHT, The 8oienoe of 8mell, (Baeia Booka, N . Y. , 1984) .
17.
R . J. BAKER, R. L . BERDEL and G . D. WALKER, E~erientla 29 . 418 (1973) .
18.
R. E . DOOLITTLE, M. BEROZA, I . KEIBER and E . L. SCHNEIDER, J . fiseot eiol. 14, 1897 (1973) .
19.
M . 8 . BLUM, R. E . DOOLTITLE and M . BEROZA, J, mseot P 2361 (1971) .
20.
L. M. RIDDII+'ORD, J. >hseot
eiol . 18, 863 (1970) .
siol . 17,