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Behavior of Japanese tree frogs under microgravity on MIR and in parabolic flight
Adv.SpaceRes.Vol.14, No. 8. pp. (8)419-(8)422,1994 Copyright@1994COSPAR Printedin GreatBritain.Allfightsreserved. 0273-1177/94$6.00+ 0.00
Pergamon
BEHAVIOR OF JAPANESE TREE FROGS UNDER MICROGRAVITY ON MIR AND IN PARABOLIC FLIGHT A. Izumi-Kurotani, 1 M. Yamashita, l Y. Kawasaki, 2 T. Kurotani, 3 Y. Mogami, 4 M. Okuno, 5 A. Oketa, 6 A. Shiraishi, 7 K. Ueda, s R. J. Wassersug 9 and T. Naitoh 10
1 Space UtilizationResearch Center, Institute of Space and Astronautical Science, Yoshino-dai, Sagamihara, Kanagawa 229, Japan 2 Laboratory of Biopolymer ConformationAnalysis, Mitsubishi Kasei Institute of Life Sciences, Japan 3 Department of Physiology, Kyoto Prefectural University of Medicine, Japan 4 Department of Biology, Faculty of Science, Ochanomizu University, Japan 5 Department of Biology, Faculty of General Education, Universityof Tokyo, Japan 6 Space Project, Tokyo Broadcasting System Inc., Japan 7 Fujitu Limited, Japan s Department of Biology, Kyoritsu University, Japan 9 Department of Anatomy and Neurobiology, Faculty of Medicine, Dalhousie University, Canada 10Department of Biology, Faculty of Science, Shimane University, Japan ABSTRACT Japanese tree frogs (Hyla japonica) were flown to the space station MIR and spent eight days in orbit during December, 1990/1/. Under microgravity, their postures and behaviors were observed and recorded. On the MIR, floating frogs stretched four legs out, bent their bodies backward and expanded their abdomens. Frogs on a surface often bent their neck backward and walked backwards. This behavior was observed on parabolic flights and resembles the retching behavior of sick frogs on land- a possible indicator of motion sickness. Observations on MIR were carded out twice to investigate the frog's adaptation to space. The frequency of failure in landing after a jump decreased in the second observation period. After the frogs retumed to earth, readaptation processes were observed. The frogs behaved normally as early as 2.5 hours after landing.
INTRODUCTION Life on earth has evolved and adapted to many environments. The behavior of animals is one expression of their response to the environments of this planet. Gravity provides a dominant coordinate for the orientation behaviors of posture and movement. Tree frogs have many gravity-dependent behaviors, i.e. walking, climbing, and jumping. They experience short periods of microgravity on earth when they jump. When frogs are exposed to microgravity for a few seconds in free fall or parabolic flight, they stretch out all four legs and bend their body backwards. This posture is similar to their jumping posture/2/. Japanese tree frogs spent eight days on the space station MIR in 1990, to investigate their behavioral responses to prolonged microgravity. These frogs showed the outstretched posture just described when floating in microgravity. Frogs on a surface often bent their neck backward and walked backwards. This posture resembled part of the retching and vomiting behavior, induced by emetic drugs in frogs/3/and is indicative of motion sickness. Observation of the frogs on MIR were performed twice to study adaptation to the space environment. Their readaptation to earth was also observed. Tree frogs in this study were also exposed to parabolic flights to determine whether they showed the same unusual postures observed on the MIR. In addition they were tested for motion sickness. Their retching postures support the hypothesis that the frogs on the MIR space station were afflicted with motion sickness. (8)419
A. I z u m i - K u r o t a n i et al.
(8)420
EXPERIMENTAL SYSTEM AND OPERATION Space Experiment Tree frogs. One hundred Japanese tree frogs, Hyla japonica, field-caught in the Kanto area of Japan, were transported to the launch site 10 days before the launch date. The frogs were starved for 10 days before the launch. In orbit, frogs were not fed routinely, but mealworms brought for an experiment to test feeding behavior were not isolated from the frogs. Twelve Japanese tree frogs (6 males and 6 females, 7-12 months old, 2 - 3 g weight) were selected for flight by several criteria: no injuries, health condition expressed by posture, and normality of vestibular functions. They were divided into 2 groups; an experimental group on MIR and a control group on the ground. Exoerimental System and operation. Outline of operation is shown in Fig. 1. Experimental system and operation are described in detail elsewhere/I,4/. parabolic Flight Ext~riment ~LC.g._fr_9~ Two kinds of tree frogs, Hyla japonica (17 specimens for flights, 22 specimens for the controls) and Rhacophorus schlegelii (24 specimens for flights, 13 for the controls) for this experiment were caught in the field, Shimane prefecture, Japan. Both frogs for flight and the controls were forcefed small pieces of beef liver 1.5-4 hours before flight. Exoerimental procedure. Specimens were installed in 2.1 liter containers with wet walls. These containers were fixed in the cabin of an MU-300 aircraft (Diamond Air Service, Nagoya, Japan). Specimens were observed during and after the flight for signs of motion sickness and recorded by 8 mm VTR camera. Retching behavior and vomiting were used as the indicators of motion sickness. The number of frogs which vomited was estimated by the number of pieces of vomitus when the vomiting occurred without observation or recording. Parabolic flight. The MU-300 flew 8-10 parabolas in one hour every day for a total of four days. About 20 seconds of microgravity were obtained in each parabola between about 10 seconds of hypergravity (1.5-2.3 G) periods.
Glove Bag
LSB SOYUZ ~ Iql
{~RF~l)very Capusule Glove Bag 1~14--FRB : Frog Recovery Box v.,~ , . 3.5 days
LSB : Life Support Box 2.5 days
i•
1st Exp. L + 3 days 2rid Exp. L + 6 days
~
LSB
LSB
~ FRB
L - 7 hrs
Launch
Recovery
Late Access
~ 1 day
FRB
R + (~ lh) R + (2 ~ 2.5hrs) Earth,Access
Fig. 1 Outline of Operation
RESULTS AND DISCUSSION Soace Exoeriment Exoeriment in orbit. Floating frogs showed a similar posture to that of frogs under short-term microgravity (i.e. free fall or parabolic flight) (Fig. 2). Both synchronous (like breaststroke swimming)
JapaneseTreeFrogBehaviorin Space
(8)421
and asynchronous movements of right and left hindlimbs were observed. The floating frogs tried to grasp any surface that their legs touched. Sometimes they rotated around their rostral-caudal axes by moving legs and/or body. The rotation was not noticed clearly under short-term microgravity. More than half (at least 4 of 6) the frogs bent their neck backward to almost right angles and pressed their abdomens against the substrate (Fig. 3). Their hindlimbs were not folded completely. In this spacespecific posture, they walked only backwards, never forward. A mission operator observed mouth opening and closing behavior, and closing of the eyes/5/. These behaviors may indicate increased salivation or swallowing. The frogs showed an avoidance response to large objects, such as the hand of a mission operator. When a mission operator brought a mealworm toward a frog on the surface of a glove bag, the frog tried to grasp it but failed to eat it. The frogs' responses to moving small objects, such as tweezers, was similar to that normally used when orienting toward such items. Orienting behavior on a rotating substance, such as a water tube, was observed. These response behaviors were similar to those shown on earth. Frequency of failure to land on a surface after jumping was counted in the VTR images. It decreased in the second observation period (i.e. an average of 2.7 random contacts with objects or surface per jump before proper landing in the first period, versus an average of 1.4 contacts per jump before landing in the second period). This result suggests that frogs could adapt to microgravity.
Fig. 2 A floating frog on the MIR space station
Fig. 3 A frog on the surface of a tool band on the MIR space station
Control exoeriment on grg0n~l. Behaviors and vestibular functions were normal for the control frogs. Behaviors of recovered frogs. All 6 frogs were recovered alive after 8 days of space flight. Observations were started at landing +2 hrs. At that time, the frogs walked and climbed slowly. Retraction of hindlimbs was abnormally delayed when they landed after jumping. Vestibular functions also did not appear to be normal. Vestibular function was assessed with three tests: Optokinetic nystagmus: Head rotation or body reorientation responses were observed when the plate, on which a frog sat, turned on its dorso-ventral axis, with an appropriate angular velocity of 20-45 degrees/sec. Head reorientation response: Reorientation of the head was observed when the plate, on which a frog sat, was tilted with an appropriate angle of 30-45 degrees from horizontal. Body righting response: This response was observed after a frog was made to lie on its back by an operator. Abnormalities in behaviors and vestibular functions disappeared within 30 minutes (at landing+2.5 hours) and were normal at landing +12 hrs. Parabolic Flight Exoeriment Under weightless conditions during the parabolic flight, more than half the frogs showed a similar posture to that shown on the MIR (Fig. 4). No specimens vomited during the parabolic flights.
(8)422
A. Izumi-Kurotani et al.
However, at least one of 17 specimens of Hyla japonica and at least four of 24 specimens of Rhacophorus schlegelii vomited after their flights. We also observed the retching behavior or mouth opening-closing behavior in some specimens during or shortly after parabolic flight. Neither vomiting nor retching behaviors were observed in the control specimens. These results indicate that frogs can get motion sickness and suggest that the unusual posture of the frogs on the MIR may have represented motion sickness.
Fig. 4 Frogs during a parabolic flight
ACKNOWLEDGMENT We thank the people listed in a report/1/, especially Mr. Toyohiro Akiyama, the first Japanese cosmonaut who executed the frog experiment on MIR in December 1990. The space experiment on MIR was supported by a research contact between the Institute of Space and Astronautical Science, and Tokyo Broadcasting System Incorporation. The flight opportunity on the MU-300 (Diamond Air Service, Japan) was funded by the National Space Development Agency of Japan. The research of Izumi-Kurotani, Yamashita, and Naltoh is supported by the Japanese Ministry of Education, Science and Culture. In addition, Izumi-Kurotani and Yamashita are supported by the Fund for Basic Experiments Oriented to Space Station Utilization of the Institute of Space and Astronautical Science. Wassersug's participation in the parabolic flight experiment was made possible by grants from the National Science and Engineering Research Council of Canada, the Japan Science and Technology Fund (External Affairs and International Trade, Canada), and contractual support from the Canadian Space Agency.
REFERENCES 1.FRIS Experiment Group, Report of Frog Experiment Onboard Space Station MIR. ed. Space Utilization Research Center, Institute of Space and Astronautical Science, Kanagawa, Japan (1991). 2.A. Izumi-Kurotani, M. Yamashita, Y. Kawasaki, T. Kurotani, Y. Mogami, M. Okuno, A. Oketa, A. Shiraishi, and K. Ueda, Behavior of Japanese Tree Frog under Microgravity. Biological Sciences in Space, 5, 185-189, (1991). 3.T. Naltoh, R. J. Wassersug, and R. A. Leslie, The Physiology, Morphology, and Ontogeny of Emetic Behavior in Anuran Amphibians. Physiological Zoology, 62, 819-843, (1989). 4.A. Izumi-Kurotani, M. Yamashita and Y. Kawasaki, Space Experiment on Behaviors of Treefrog. Adv. Space Res. 12, # 1,263 (1992). 5.T. Akiyama, private communication (1991).
Pergamon
BEHAVIOR OF JAPANESE TREE FROGS UNDER MICROGRAVITY ON MIR AND IN PARABOLIC FLIGHT A. Izumi-Kurotani, 1 M. Yamashita, l Y. Kawasaki, 2 T. Kurotani, 3 Y. Mogami, 4 M. Okuno, 5 A. Oketa, 6 A. Shiraishi, 7 K. Ueda, s R. J. Wassersug 9 and T. Naitoh 10
1 Space UtilizationResearch Center, Institute of Space and Astronautical Science, Yoshino-dai, Sagamihara, Kanagawa 229, Japan 2 Laboratory of Biopolymer ConformationAnalysis, Mitsubishi Kasei Institute of Life Sciences, Japan 3 Department of Physiology, Kyoto Prefectural University of Medicine, Japan 4 Department of Biology, Faculty of Science, Ochanomizu University, Japan 5 Department of Biology, Faculty of General Education, Universityof Tokyo, Japan 6 Space Project, Tokyo Broadcasting System Inc., Japan 7 Fujitu Limited, Japan s Department of Biology, Kyoritsu University, Japan 9 Department of Anatomy and Neurobiology, Faculty of Medicine, Dalhousie University, Canada 10Department of Biology, Faculty of Science, Shimane University, Japan ABSTRACT Japanese tree frogs (Hyla japonica) were flown to the space station MIR and spent eight days in orbit during December, 1990/1/. Under microgravity, their postures and behaviors were observed and recorded. On the MIR, floating frogs stretched four legs out, bent their bodies backward and expanded their abdomens. Frogs on a surface often bent their neck backward and walked backwards. This behavior was observed on parabolic flights and resembles the retching behavior of sick frogs on land- a possible indicator of motion sickness. Observations on MIR were carded out twice to investigate the frog's adaptation to space. The frequency of failure in landing after a jump decreased in the second observation period. After the frogs retumed to earth, readaptation processes were observed. The frogs behaved normally as early as 2.5 hours after landing.
INTRODUCTION Life on earth has evolved and adapted to many environments. The behavior of animals is one expression of their response to the environments of this planet. Gravity provides a dominant coordinate for the orientation behaviors of posture and movement. Tree frogs have many gravity-dependent behaviors, i.e. walking, climbing, and jumping. They experience short periods of microgravity on earth when they jump. When frogs are exposed to microgravity for a few seconds in free fall or parabolic flight, they stretch out all four legs and bend their body backwards. This posture is similar to their jumping posture/2/. Japanese tree frogs spent eight days on the space station MIR in 1990, to investigate their behavioral responses to prolonged microgravity. These frogs showed the outstretched posture just described when floating in microgravity. Frogs on a surface often bent their neck backward and walked backwards. This posture resembled part of the retching and vomiting behavior, induced by emetic drugs in frogs/3/and is indicative of motion sickness. Observation of the frogs on MIR were performed twice to study adaptation to the space environment. Their readaptation to earth was also observed. Tree frogs in this study were also exposed to parabolic flights to determine whether they showed the same unusual postures observed on the MIR. In addition they were tested for motion sickness. Their retching postures support the hypothesis that the frogs on the MIR space station were afflicted with motion sickness. (8)419
A. I z u m i - K u r o t a n i et al.
(8)420
EXPERIMENTAL SYSTEM AND OPERATION Space Experiment Tree frogs. One hundred Japanese tree frogs, Hyla japonica, field-caught in the Kanto area of Japan, were transported to the launch site 10 days before the launch date. The frogs were starved for 10 days before the launch. In orbit, frogs were not fed routinely, but mealworms brought for an experiment to test feeding behavior were not isolated from the frogs. Twelve Japanese tree frogs (6 males and 6 females, 7-12 months old, 2 - 3 g weight) were selected for flight by several criteria: no injuries, health condition expressed by posture, and normality of vestibular functions. They were divided into 2 groups; an experimental group on MIR and a control group on the ground. Exoerimental System and operation. Outline of operation is shown in Fig. 1. Experimental system and operation are described in detail elsewhere/I,4/. parabolic Flight Ext~riment ~LC.g._fr_9~ Two kinds of tree frogs, Hyla japonica (17 specimens for flights, 22 specimens for the controls) and Rhacophorus schlegelii (24 specimens for flights, 13 for the controls) for this experiment were caught in the field, Shimane prefecture, Japan. Both frogs for flight and the controls were forcefed small pieces of beef liver 1.5-4 hours before flight. Exoerimental procedure. Specimens were installed in 2.1 liter containers with wet walls. These containers were fixed in the cabin of an MU-300 aircraft (Diamond Air Service, Nagoya, Japan). Specimens were observed during and after the flight for signs of motion sickness and recorded by 8 mm VTR camera. Retching behavior and vomiting were used as the indicators of motion sickness. The number of frogs which vomited was estimated by the number of pieces of vomitus when the vomiting occurred without observation or recording. Parabolic flight. The MU-300 flew 8-10 parabolas in one hour every day for a total of four days. About 20 seconds of microgravity were obtained in each parabola between about 10 seconds of hypergravity (1.5-2.3 G) periods.
Glove Bag
LSB SOYUZ ~ Iql
{~RF~l)very Capusule Glove Bag 1~14--FRB : Frog Recovery Box v.,~ , . 3.5 days
LSB : Life Support Box 2.5 days
i•
1st Exp. L + 3 days 2rid Exp. L + 6 days
~
LSB
LSB
~ FRB
L - 7 hrs
Launch
Recovery
Late Access
~ 1 day
FRB
R + (~ lh) R + (2 ~ 2.5hrs) Earth,Access
Fig. 1 Outline of Operation
RESULTS AND DISCUSSION Soace Exoeriment Exoeriment in orbit. Floating frogs showed a similar posture to that of frogs under short-term microgravity (i.e. free fall or parabolic flight) (Fig. 2). Both synchronous (like breaststroke swimming)
JapaneseTreeFrogBehaviorin Space
(8)421
and asynchronous movements of right and left hindlimbs were observed. The floating frogs tried to grasp any surface that their legs touched. Sometimes they rotated around their rostral-caudal axes by moving legs and/or body. The rotation was not noticed clearly under short-term microgravity. More than half (at least 4 of 6) the frogs bent their neck backward to almost right angles and pressed their abdomens against the substrate (Fig. 3). Their hindlimbs were not folded completely. In this spacespecific posture, they walked only backwards, never forward. A mission operator observed mouth opening and closing behavior, and closing of the eyes/5/. These behaviors may indicate increased salivation or swallowing. The frogs showed an avoidance response to large objects, such as the hand of a mission operator. When a mission operator brought a mealworm toward a frog on the surface of a glove bag, the frog tried to grasp it but failed to eat it. The frogs' responses to moving small objects, such as tweezers, was similar to that normally used when orienting toward such items. Orienting behavior on a rotating substance, such as a water tube, was observed. These response behaviors were similar to those shown on earth. Frequency of failure to land on a surface after jumping was counted in the VTR images. It decreased in the second observation period (i.e. an average of 2.7 random contacts with objects or surface per jump before proper landing in the first period, versus an average of 1.4 contacts per jump before landing in the second period). This result suggests that frogs could adapt to microgravity.
Fig. 2 A floating frog on the MIR space station
Fig. 3 A frog on the surface of a tool band on the MIR space station
Control exoeriment on grg0n~l. Behaviors and vestibular functions were normal for the control frogs. Behaviors of recovered frogs. All 6 frogs were recovered alive after 8 days of space flight. Observations were started at landing +2 hrs. At that time, the frogs walked and climbed slowly. Retraction of hindlimbs was abnormally delayed when they landed after jumping. Vestibular functions also did not appear to be normal. Vestibular function was assessed with three tests: Optokinetic nystagmus: Head rotation or body reorientation responses were observed when the plate, on which a frog sat, turned on its dorso-ventral axis, with an appropriate angular velocity of 20-45 degrees/sec. Head reorientation response: Reorientation of the head was observed when the plate, on which a frog sat, was tilted with an appropriate angle of 30-45 degrees from horizontal. Body righting response: This response was observed after a frog was made to lie on its back by an operator. Abnormalities in behaviors and vestibular functions disappeared within 30 minutes (at landing+2.5 hours) and were normal at landing +12 hrs. Parabolic Flight Exoeriment Under weightless conditions during the parabolic flight, more than half the frogs showed a similar posture to that shown on the MIR (Fig. 4). No specimens vomited during the parabolic flights.
(8)422
A. Izumi-Kurotani et al.
However, at least one of 17 specimens of Hyla japonica and at least four of 24 specimens of Rhacophorus schlegelii vomited after their flights. We also observed the retching behavior or mouth opening-closing behavior in some specimens during or shortly after parabolic flight. Neither vomiting nor retching behaviors were observed in the control specimens. These results indicate that frogs can get motion sickness and suggest that the unusual posture of the frogs on the MIR may have represented motion sickness.
Fig. 4 Frogs during a parabolic flight
ACKNOWLEDGMENT We thank the people listed in a report/1/, especially Mr. Toyohiro Akiyama, the first Japanese cosmonaut who executed the frog experiment on MIR in December 1990. The space experiment on MIR was supported by a research contact between the Institute of Space and Astronautical Science, and Tokyo Broadcasting System Incorporation. The flight opportunity on the MU-300 (Diamond Air Service, Japan) was funded by the National Space Development Agency of Japan. The research of Izumi-Kurotani, Yamashita, and Naltoh is supported by the Japanese Ministry of Education, Science and Culture. In addition, Izumi-Kurotani and Yamashita are supported by the Fund for Basic Experiments Oriented to Space Station Utilization of the Institute of Space and Astronautical Science. Wassersug's participation in the parabolic flight experiment was made possible by grants from the National Science and Engineering Research Council of Canada, the Japan Science and Technology Fund (External Affairs and International Trade, Canada), and contractual support from the Canadian Space Agency.
REFERENCES 1.FRIS Experiment Group, Report of Frog Experiment Onboard Space Station MIR. ed. Space Utilization Research Center, Institute of Space and Astronautical Science, Kanagawa, Japan (1991). 2.A. Izumi-Kurotani, M. Yamashita, Y. Kawasaki, T. Kurotani, Y. Mogami, M. Okuno, A. Oketa, A. Shiraishi, and K. Ueda, Behavior of Japanese Tree Frog under Microgravity. Biological Sciences in Space, 5, 185-189, (1991). 3.T. Naltoh, R. J. Wassersug, and R. A. Leslie, The Physiology, Morphology, and Ontogeny of Emetic Behavior in Anuran Amphibians. Physiological Zoology, 62, 819-843, (1989). 4.A. Izumi-Kurotani, M. Yamashita and Y. Kawasaki, Space Experiment on Behaviors of Treefrog. Adv. Space Res. 12, # 1,263 (1992). 5.T. Akiyama, private communication (1991).