Space environment effect on circadian rhythms of plants for use in biosatellit

French, J. D.

Hoshizaki, T.

English

NASA Technical Reports Server

,I I \ 
I\ 
FINAL REPORT TO 
NATIONAL AERONAUTICS AND SPACE ADMINISTRATION 
Research Grant NsG - 528 
E f f e c t  o f  Space Envi ronment on C i rcad ian Rhythms 
o f  Plants,  For the Purpose of  De f in ing  and V e r i f y i n g  an 
Experiment Su i tab le  f o r  Use i n  a B i o s a t e l l i t e .  
September 1, 1964 t o  February 28, 1965 
D r ,  J. D. French 
D r .  T. Hoshizaki 
Bra in  Research I n s t i t u t e  
U n i v e r s i t y  o f  Cal i f o t n i a ,  Los Angeles 
I I ., 4 N 6 5 - 2 3 7 4 2  UCCESSIO UMBER) - 
I 
> IPAGES) 57 
(37429477 
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(NASA C R M  f i x  OR AD NUhfBER) ICATEG 
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February 28, 1965 GPO PRICE $ - 
OTS PRICE(S) $ 
https://ntrs.nasa.gov/search.jsp?R=19650014141 2020-03-17T00:05:39+00:00Z
Fina l  Report 
Grant NsG 528 
This  I s  the f i n a l  repor t  o f  research performed under NASA 
~ 
Grant 528-63, f o r  the purpose of  de f fn lng  and v e r i f y i n g  an experiment 
su i tab le  fo r  use i n  a B i o s a t e l l i t e .  The experiment i s  t o  t e s t  the 
e f f e c t  o f  space environment on the c i r cad ian  rhythms o f  p lan ts :  
lea f  movements o f  the P in to  bean p lan t  (Phaseolus v u l g a r i s )  arid 
growth bands o f  fuvgus (Neurospora crassa 2!863) 2 3 7 y v  
-- Our work has resu i ted  i n  (1)  the development o f  the MOGRAVATRON, 
an apparatus t o  slrnulate we?ghtlessness cond l t l on  f o r  p lan ts ,  (2) 
the development o f  an ln f ra - red  time lapse photographic system, ( 3 )  
the develo7ment o f  a s t r a l n  gauge recording system whlch measures 
the movements o f  P i n t o  leaves and whose s!gnal Is su i tab le  f o r  
te lemetry,  (4) the development o f  prototype f l  Igh t  package f o r  
Neurospora experiments, (5) f i nd ing  response d f f fe rences  I n  growth 
rates and growth d i r e c t i o n s  I n  Bar ley seedl ings when they were 
grown i n  a simulated weightlessness c m d t t i o n ,  (6) conf i rmat ion  and 
de l i nea t ion  o f  the l e a f  movement when the p l a n t  i s  s ta t i ona ry  and 
grown under constant condi t ions,  (7) the reduct ion i n . t b  s ize  o f  the 
experfmental p lan t  mater la l ,  (8) f i n d i n g  t h a t  the l e a f  movements o f  
P i n t o  beans respond t o  a simulated weightlessness condf t ian  by the 
loss o f  l e a f  movements o r  by a phase s h i f t  o f  the l e a f  movements, 
(9) conf i rmat ion o f  the bas ic  27 hour l e a f  movement rhythm by ccm- 
p u t c r  ana lys is  o f  photographic t ime lapse data. 
/-The e a r l y  r e s u l t s  which have been presented i n  the previous 
repor ts  w i l l  be summarized In t h i s  repor t  and r e s u l t s  t h a t  were 
obtalned i n  the recent experiments w i l l  be discussed i n  some d e t a i l .  
The r e s u l t s  t o  be reported are f rom experimental work done I n  the 
Space Bio logy Laboratory a t  the Un lve rs l t y  o f  C a l  i f o r n l a ,  Los Angeles 
and a t  the  Ames Research Center, Mof fe t t  F le ld ,  Ca l i  fo rn ia .  
work i s  an t i c ipa ted  f o r  the  l a t e s t  ser ies  o f  experlments before the 
p u b l i c a t i o n  o f  the f ind ings.  
Fur ther  
DEVELOPMENT OF SPECIAL APPARATUS 
During the i n i t i a l  s i x  month per iod o f  the grant,  most o f  
the e f f o r t  was d i r e c t e d  toward the development o f  the experimental 
apparatus and the s e l e c t i o n  o f  the appropr ia te experimental methods 
and tk refinement o f  the experimental techniques. 
The Nogravatron . A Weightlessness Simulator. 
The Nogravatron was developed and the simulated weightlessness 
envircnment was tested by growing A t l a s  b a r l e y  seedl ings on the 
Mogravatrcn. The simulated weightlessness c o n d i t i o n  i s  achieved by 
r o t a t i n g  the p l a n t  simultaneously about two axes perpendicular t o  
each other.  The p l a n t  i s  ro ta ted  a t  the r a t e  o f  one r e v o l u t i o n  per 
minute around one a x i s  and one revo lu t ion  i n  four  minutes around the 
second ax is .  
Time Lapse Photographic System 
The time lapse 16 mm camera and accessory equipment were 
adapted t o  the Nogravatron t o  expose one frame every t ime the ro ta ted  
p l a n t s  reached an u p r i g h t  pos i t ion .  In  order  t o  take p i c t u r e s  
dur ing  both the dark phase and the l i g h t  phase i n  the var ious e x p e r i -  
mental regimes used i n  our experiments, spec ia l  i n f r a - r e d  lamps were 
designed, fabr ica ted  and adapted f o r  t h i s  p a r t i c u l a r  purpose. The 
in f ra - red  photographic record obtained by t h i s  system were o f  h i g h  
q u a l i t y .  
S t r a i n  Gauge Leaf Movement Recording System 
In  a d d i t i o n  t o  the time lapse photographic method, a s t r a i n  
gauge l e a f  movement recording system was developed as an a l t e r n a t i v e  
te lemetry  system. This  system which i s  small and l i g h t  weight con- 
s i s t s  o f  a m i c r o - s t r a i n  gauge glued t o  a brass shim stock measuring 
I , , / 1 r  in& wide ‘ jy  ; 1 / I .  I - -L  l r r -n  
I / ’ +  IIILII I”1’y. T h i s  s h i m  stock I s  g !W!  tc! t h e  
blade and t o  the p e t i o l e .  The s t r a i n  r e s u l t i n g  from the l e a f  move- 
ment i s  amp1 i f i e d  and recorded by appropr ia te instruments. 
' .  . . '  
-3 - 
Prototype F1 i g h t  Package f o r  Neurospora crassa 21853 
I n  a d d i t i o n  t o  the development o f  ground based experimental 
equipment, a two pound prototype f l i g h t  package was developed a t  the 
Space B ic logy  Laboratory f o r  o r b i t a l  f l i g h t .  This u n i t  was f l i g h t  
stressed t o  73 g ' s  a t  2 KC. 
c o n t r o l s  and the Neurospora fungi  were no t  damaged by these condi t ions.  
The subsequent growth o f  the Neurospora fungi  appeared normal. I n  
l a t e r  experiments, v a r i a t i o n  i n  the growth p a t t e r n  was noted and 
f u r t h e r  s tud ies showed t h a t  these v a r i a t i o n s  could n o t  be e l im ina ted  
by standard i s o l a t i o n  techniques. Due t o  t h i s  v a r i a t i o n  problem i n  
the Neurospora stock c u l t u r e  and due t o  the shortage o f  h e l p  and 
time, the Neurospora experiment was set aside f o r  the t ime being and 
our e f f o r t s  were placed on the l e a f  movement experiments, 
The package w i t h  i t s  environmental 
S I MULATED WE IGHTLESSNESS STUD 1 ES 
US I NG ATLAS BARLEY SEEDL I NGS 
To t e s t  the simulated weightlessness condi t ions created by the 
2 axes r o t a t i o n s  o f  the Nogravatron, bar ley  seedl ings were germinated 
and grown on the apparatus, The Nogravatron grown seedl ings i n i t i a l l y  
grew f a s t e r  than the s t a t i o n a r y  cont ro l  seedl ings i n d i c a t i n g  a growth 
s t i m u l a t i o n  e f f e c t  o f  the simulated weightlessness environment. The 
growth d i r e c t i o n  o f  the Nogravatron grown shoot i s  determined by the 
o r i e n t a t i o n  o f  the embryo. Upon germination i n  the s imulated 
weightlessness environment, the shoot w i l l  grow out  f o l l o w i n g  a l i n e  
co inc ident  w i t h  the d i r e c t i o n  o f  the o r i g i n a l  embryo shoot s t ruc tu re .  
On the  o ther  hand, the growth d i r e c t i o n  o f  the roo ts  appears t o  depend 
on the i n i t i a l  s p a t i a l  r e l a t i o n s h i p  i.e. a crowding e f f e c t  t h a t  develops 
as the  roots  a r e  emerging from the seed and expanding i n  g l k t h .  Thus 
the Nogravatron does indeed produce an ageotropic c o n d i t i o n  for the 
Bar ley seedi ings. 
-4 - 
PINTO BEAM LEAF MOVEMENT STUDIES 
Conf i rmatory t e s t s  
I n  the e a r l y  p a r t  o f  t h i s  grant, l e a f  movement experiments were 
run a t  the Ames Research Laboratory i n  order  t o  determine the 
environmental l i m i t s  w i t h i n  which the P i n t o  bean would cont inue t o  
e x h i b i t  i t s  l e a f  movemmts. A t  the same t ime these experiments 
confirmed the e a r l  i e r  l e a f  movement s tud ies and es tab l  ished f i rmer 
base l i n e  data f o r  P i c t o  bean leaf  movements. 
Reduction o f  Experimental P l a n t  Size 
The s i z e  o f  the experimental materi  a1 was subs tan t ia l  l y  
reduced due t o  the work done a t  the Ames Research Laboratory, The 
p l a n t  l e a f  mater ia l  has been reduced to a p e t i o l e  and l e a f  m i d r i b  
both o f  which are 1 cm i n  length. These very  reduced p l a n t  mater ia ls  
s t i l  e x h i b i t  c i rcad ian  l e a f  movements. A repor t  on t h i s  phase o f  
work was made a t  the I964 American I n s t i t u t e  o f  B i o l o g i c a l  Sciences 
meet ng h e l d  a t  Boulder, Colorado. 
Simulated Weightlessness E f f e c t s  on P i n t o  Bean Leaf Movements 
P i n t o  bean p lan ts  were grown i n  the green house u n t i l  the 
pr imary leaves were we l l  developed, These p l a n t s  were then placed i n  
a growth chamber w i t h  constant temperature and continuous 1 i gh t .  
Four p l a n t s  were used as s ta t ionary  c o n t r o l s  and f o u r  p l a n t s  were 
placed on the Nagravetron. Time lapse movie records were taken f o r  
three days dur ing  the p r e - r o t a t i o n  per iod  f o r  base l i n e  data. A t  the 
beginning o f  the f o u r t h  day, the Nogravatron was s t a r t e d  and the 
f o u r  mounted p l a n t s  began t h e i r  simulated weightlessness existence. 
Time lapse movie records were obtained dur ing  the whole course o f  
the  experiment which consisted o f  3 s t a t i o n a r y  days fol lowed by 7 
A - s a -  - C  . - t . - . , l - & - J  ..-:-h*l------e ennJ:+:nn- u a y a  V I  J IIIIU I a ccju w c  I yi ir  1 ~ : 5 a i i c a a  LUIIU I L I UIIJ.  
P I N T O  BEAN L E A F  MOVEMENT AS AFFECTED BY THE ONSET O F  
R O T A T I O N  A T  D I F F E R E N T  PHASES O F  THE C I R C A D I A N  RHYTHM 
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ROTAT I ON 
-- S T A T  I ONARY - ROTATED 
I I I I I I I I I I I 
I I I I I I I I 1 I 1 
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-5 - 
Pre l  iminary analys is  o f  the data (Fig. 1 ,) ind icates t h a t  i f  
r o t a t i o n  i s  s ta r ted  when the leaves are i n  t h e i r  opening o r  morning 
phase, the leaves appear t o  lose t h e i r  rhythmic movements and may 
not  recover t h e i r  rhy thmic i t y  f o r  as long a s  60 hours. 
the leaves were found t o  be i n  synchrony w i t h  the normal s ta t i ona ry  
con t ro l  leaves. ‘if the r o t a t i o n  i s  i n i t i a t e d  dur ing the e a r l y  
c los ing  o r  the af ternoon phase, there i s  an immediate s h i f t  i n  the 
rhythmic p a t t e r n  o f  the leaves and the leaves do no t  appear t o  lose 
t h e i r  rhythmic movements. 
Upon recovery, 
It appears t h a t  a change i n  the  g r a v i t a t i o n a l  environment has 
the same e f f e c t  on the c i rcad ian  rhythm a s  those o f  l i g h t  and o f  
temperature. Further experiments are p resent ly  being conducted t o  
look more c lose ly  a t  t h i s  p a r t i c u l a r  response. 
Computer Analys is  o f  Leaf Movement Da ta  
With the cooperat ion o f  the Heal th  Science Computing Centtasl. 
a t  UCLA, ana lys is  o f  bean l e a f  movement data have been made. From 
the data generated by analys is ,  the P in to  bean leaves have a bas ic  27 
hour p e r i o d i c i t y  i n  t h e i r  l e a f  movements. Other rhythms o f  shor te r  
p e r i o d i c i t y  do not appear t o  be present i n  leaves grown under constant 
condi t ions.  
sence o f  o the r  rhythms shor te r  o r  longer than the 27 hour rhythm 
which has been proven t o  be so prevalent .  
be made on l e a f  movement data obta ined from p lan ts  grown under a 
s imulated weightlessness condi t ion.  
Fur ther  analyses are being made t o  determine the p re -  
I n  add i t ion ,  ana lys is  w i l l  
ACKNOWLEDGEMENTS 
We wiJh t o  thank a t  t h i s  time the gran t ing  agency, the Nat ional  
Aeronautics and Space Admin is t ra t ion  f o r  t h e i r  generos i ty  i n  support ing 
t h i s  research. We a l s o  wish t o  thank the many ind i v idua l  members o f  
NASA f o r  t h e i r  e f f o r t  and cooperation i n  making i t  poss ib le  f o r  us 
t o  do our work. 
Research B i o s a t e l l i t e  group go our specia l  thanks. Without t h e i r  co- 
operat ion and help, pa r t s  o f  t h i s  work would not have been possible.  
To Drs. K. Yokoyama and la!, H. Jones o f  the Ames 
Reports 
French, J. D. and T. Hoshizaki.  Semi-annual Report f o r  the per iod  
ending March 1, 1964. E f f e c t  o f  Space Environment on Ci rcadian 
Rhythms o f  Plants,  For the Purpose o f  Def in ing  and V e r i f y i n g  an 
Experiment Su i tab le  f o r  Use i n  a B i o s a t e l l i t e .  
French, J. D. and T. Hoshizaki.  Semi-annual Report f o r  the per iod  
ending September 1, 1964. E f f e c t  o f  Space Environment on 
Ci rcadian RhycSms o f  Plants, For t h e  Furpose o f  D e f i n i n g  and 
V e r i f y i r g  an  Experiment Sui tab le f o r  Use in a B i o s a t e l l i t e .  
Pub l ica t ions  
Hoshizaki,  T. Yokoyama, K., and Jones, W .  H .  Observations o f  the 
Rhythmic Movements o f  Excised Phaseolus Leaf. P lan t  Physiology 
39 (supp .) X X X V l  i I  
Hoshizaki,  T., and Yokoyama, K. Recording Leaf Movements w i t h  a 
S t r a i n  Gauge (Submitted f o r  pub1 i c a t i o n  t o  Nature) 
Hoshizaki,  T. E f f e c t s  o f  Rotation Around Two Axes on the Growth o f  
A t l a s  Bar ley Seed1 ings. ( i n  manuscript form) 


Effect of space environment on circadian rhythms of plants, for the purpose of defining and verifying an experiment suitable for use in a biosatellite  final report, sep. 1, 1964 - feb. 28, 1965

Effect of space environment on circadian rhythms of plants, for the purpose of defining and verifying an experiment suitable for use in a biosatellite final report, sep. 1, 1964 - feb. 28, 1965

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