Experiment DCL(SLS-1) 1/1

File Name: DC85.DOC

Effects of Microgravity-induced Weightlessness on [Aurelia]{.underline}
Differentiation and Statolith Synthesis.[^1]\*

Dorothy B. Spangenberg, Principal Investigator

Jellyfish were flown in space for the first time aboard SLS-1 on June 5,
1991 for a period of nine days. Because it was the first flight for
developing jellyfish, the jellyfish experiment was designed to
facilitate ephyra formation by flying groups of animals induced by
iodine or thyroxine in space and at intervals prior to launch.
Thirty-two groups representing a total of 2,478 jellyfish were
maintained in the mid-deck locker during the launch, transferred to 28°C
for eight days, and returned to the mid-deck locker the day before
re-entry. Six groups of Earth-developed ephyrae were flown for behavior
studies and twenty-six groups (Kit 1 and 2) were designated for
developmental studies (Table 1). Of the latter groups, six were fixed in
space with glutaraldehyde on day 8. Six groups were induced to
strobilate (metamorhose) in space on Day 1. Four of these groups and two
groups induced on Earth prior to flight (48h) were fixed in space with
glutaraldehyde on day 8. Other polyp groups were induced to strobilate
either 48 or 24h prior to launch. Two of the polyp groups were not given
inducer, but were flown in artificial sea water (ASW). Six groups of
Earth-developed ephyrae were flown for behavioral studies. Thirty-two
ground-based control groups were maintained under simulated flight
conditions; fourteen of these groups were tested in the same time period
as the flight experiment animals and eighteen groups were tested one
week post-flight. (See Spangenberg, et al, 1993a).

The objectives of the experiment were: (1) to determine whether the
microgravity of outer space will modify the development of ephyrae from
polyps; the development of the graviceptors (rhopalia) of ephyrae; the
formation or demineralization of statoliths of rhopalia; or the
swimming/pulsing behavior of ephyrae. (2) By comparing the features
listed in (1) in ephyrae which developed in space with those that
developed on Earth, to discover the role that gravity plays on Earth in
the development of ephyrae, their graviceptors, and their behavior.

[Results]{.underline} [of]{.underline} [Development]{.underline}
[Studies]{.underline}: The number of ephyrae formed, the number of arms
and rhopalia/ephyra, and the number of statoliths/rhopalia was compared
between space-developed and ground-based control animals. It was found
that the average number of ephyrae formed per polyp was slightly higher
in the Earth-developed groups (2.95 ephyrae/polyp) as compared with
those that developed during the flight (2.74/polyp). Average arm numbers
were statistically significantly lower in the space-induced organisms
(8.19/ephyra) than in controls (8.64/ephyra). Since eight arms/ephyra is
the number considered most normal, the in-flight animals had a more
normal average arm number than controls. There were no significant
differences between the rhopalia numbers of the flight animals and
controls; however, statolith numbers were significantly higher
(23.18/rhopalium) in space-induced organisms than their controls
(17.94/rhopalium). Animals induced on Earth and then sent into space at
L-24h and L-48h showed no difference in the number of statoliths
synthesized as compared with those induced on Earth and retained on
Earth.

Statolith numbers were statistically significantly lower in
thyroxine-induced organisms than in iodine-induced jellyfish both in
space-developed and ground-developed ephyrae.

[Ephyra]{.underline} [Formation]{.underline} [without]{.underline}
[Inducer]{.underline}: Two groups of polyps sent into space in ASW
produced ephyrae. Earth-control animals maintained in the same batch of
ASW and exposed to comparable temperature range, did not form ephyrae.
In addition, two more controls in the same batch of ASW tested
postflight did not form ephyrae. It is know that the Jf-T4 hormone is
secreted into the medium (Spangenberg, 1971) and it is also know that
brine shrimp on which jellyfish subsist contain iodine. The flight
jellyfish were fed on the same schedule as controls. The formation of
ephyrae in replicates in ASW in space, therefore, suggests that hormone
production, utilization, or accumulation in the medium (with possible
subsequent feed-back on the organisms) was different in space animals
compared with ground-controls. If the induction response was caused by a
localized build-up of hormone in the ASW in space, then the implication
is that secretory products from cells or other aquatic

animals may accumulate differently in space and create unexpected
biological responses.

[Statolith]{.underline} [Demineralization]{.underline}: Demineralization
of statoliths in non-fed ephyrae that were flown in space in ASW as
compared to Earth-maintained ephyrae was increased. Comparison of
base-line statolith numbers from the same group of ephyrae as those used
for the space experiment revealed a 47.7% loss of statoliths in space
maintained animals as compared with a 6.3% loss of statoliths in ground
controls. Ephyrae maintained in thyroxine in space and one the had a
58.1% and 53.6% statolith loss respectively. Apparently, the thyroxine
functioned in space by increasing demineralization of statoliths as on
Earth, (Spangenberg, 1984), but there apparently was not a synergistic
effect between thyroxine and microgravity effects which would have
caused a significantly increased statolith loss in the space-maintained
ephyrae in thyroxine as compared with ground controls.

[Morphology]{.underline} [of]{.underline} [Space-developed]{.underline}
[and]{.underline} [Earth-developed]{.underline} [Ephyrae]{.underline}:

Light Microscopy: Examination of two hundred ephyrae from space and
their ground-based controls immediately post-flight with dissecting
microscope revealed no morphological differences in the space-developed
ephyrae as compared with the controls. Distribution of arms, rhopalia,
statoliths, gastric filaments, and nematocyst clusters was normal as
compared with controls. In short, the patterning of the organisms was
similar whether they developed in space or on Earth. In addition,
examination of twenty ephyrae with the SEM revealed that ciliation on
the surfaces of the ephyrae was normal in both groups of jellyfish.

Examination of the various structures of the ephyrae at the
ultrastructural levels is still in progress. To date, examination of
graviceptors (rhopalia) and radial muscle of several organisms at the
TEM (transmission electron microscope) level reveals a close similarity
between the groups. The space-developed animals had well-developed
rhopalia with touch-plates which were characteristic for this age
organism. The ultrastructural studies of rhopalia and striated muscle,
however, are still in progress.

[Behavior]{.underline} [Studies]{.underline}: Normal behavior in
newly-developed organisms is another parameter of normal development.
Ephyrae which developed in space were able to integrate their
neuro-muscular systems with their graviceptors to permit complete
pulsing contractions and consequently to achieve swimming. This swimming

behavior, however, was not constant with swimming behavior of
Earth-controls. On Earth, when ephyrae quit pulsing and stop swimming,
they sink due to the effect of gravity. In space, when the organisms
stopped pulsing the could not sink and were immobilized in place until
they began pulsing again. Many tended to swim in circles. It was
significant to learn that ephyrae were capable of swimming and pulsing
in space without the stimulus of gravity. Some organisms, however, that
developed in space had pulsing abnormalities.

[Pulsing]{.underline} [abnormalities]{.underline}: Of particular
interest were some of the ephyrae which were examined within 12h
post-flight. Some (18.1%) of the space-developed ephyrae had pulsing
abnormalities while the ground-based controls had very few pulsing
abnormalities (2.9%). Ephyrae with incomplete pulses, spasms,
aftertwitches, uncoordinated pulsing, and \"waves\" ( a circular
rippling of a pulse) were found only in the space-developed groups. One
control was found with jerky pulsing which was not seen in the
space-developed ephyrae. Pulsing activity in ephyrae is controlled by an
interaction between the rhopalia, the nerve-nets, and the neuro-muscular
systems. It will be important to determine whether the abnormal pulsing
in space-developed ephyrae reflects an abnormality in the development of
the graviceptors, the nerve nets, or the neuro-muscular system or if the
problem(s) arise from a lack of integration between these structures.

[Acknowledgments]{.underline}: This research was funded by NASA grants
NAG2-343 and NAGW-1784. The P.I. is grateful to the following people for
their assistance with the Jellyfish-in-Space Experiment: The SLS-1 crew,
especially Dr. T. Jernigan who did the space portion of the experiment;
the Eastern Virginia Medical School Jellyfish Research Team: Dr. R.
McCombs, Dr. J. Shaeffer, Dr. R. Spangenberg, B.T. Lowe, M. Sampson, J.
Slusser, A. Shore, M. Hughes, G. Shelton, C. Ramiro, D. Leete, A.
Barnes, M. McCombs, and M. Prokopchak; Dr. Philput, Naval Hospital,
Portsmouth, statistics consultant; ARC: Dr. G. Jahns, Dr. C. Winget, S.
Jones, B. Dalton, G. McCollough, D. Cheng, and T. Nelson; NASA HQ: Dr.
T. Halstead and Dr. R. White.

[References]{.underline}:

Spangenberg, D.B. 1971. Thyroxine Induced Metamorphosis in
[Aurelia]{.underline}. J. Exp. Zool. 178:183-194

Spangenberg, D.B. 1984. Effects of Exogenous Thyroxine on Statolith
Synthesis. Am. Zool. 24:917-923.

Spangenberg, D.B., Jernigan, T., McCombs, R., Lowe, B., Sampson, M. and
Slusser, J. 1993a. Developmental Studies of [Aurelia]{.underline}
(Jellyfish) Ephyrae Which Developed During the SLS-1 Mission. Adv. Space
Res., In Press.

Spangenberg, D.B., Jernigan, T., Philput, C. and Lowe, B., 1993b,
Graviceptor Development in Jellyfish Ephyrae in Space and on Earth. Adv.
Space Res., In Press.

[TABLE 1]{.underline} Space Flown Jellyfish in Kits 1 and 2

+-----------------------------------+-----------------------------------+
| 100 Polyps in 149 ml ASW induced  | 66 Polyps in 100 ml ASW induced   |
| on Earth to                       | in space to                       |
|                                   |                                   |
| metamorphose with:                | metamorphose with:                |
+-----------------------------------+-----------------------------------+

  ----------- ----------- ----------- ----------- ----------- -----------
  I2 @ L -    I2 @ L -    \*I2 @ L -  I2 @ L + 8h I2 @ L + 8h \*\*I2 @ L+
  24h         24h         24h                                 8h

  ----------- ----------- ----------- ----------- ----------- -----------

  ----------- ----------- ----------- ----------- ----------- -----------
  T4 @ L -    T4 @ L -    \*T4 @ L -  T4 @ L + 8h T4 @ L + 8h \*\*T4 @
  24h         24h         24h                                 L + 8h

  ----------- ----------- ----------- ----------- ----------- -----------

  ----------- ----------- ----------- -----------------------------------
  I2 @ L -    I2 @ L -    \*I2 @ L -  66 Polyps in 100 ml ASW induced on
  48h         48h         48h         Earth to

  ----------- ----------- ----------- -----------------------------------

  ----------- ----------- ----------- -----------------------------------
  T4 @ L -    T4 @ L -    \*T4 @ L -  metamorphose with:
  48h         48h         48h         

  ----------- ----------- ----------- -----------------------------------

  ----------------------------------- ----------- ----------- -----------
  100 Polyps in 149 ml ASW w/o I2 or  \*\*I2 @                
  T4 added:                           L - 48h                 

  ----------------------------------- ----------- ----------- -----------

  ----------- ----------- ----------- ----------- ----------- -----------
  @ L - 24h   \*@ L-24h               \*\*T4@ L -             
                                      48h                     

  ----------- ----------- ----------- ----------- ----------- -----------

\* Fixed on ground after flight

\*\*Fixed in space

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