The effect of a diminished motor activity of rats upon the ribonucleic and (RNA) content in a single isolated motoneuron of frontal of their spinal cord was studied. Within a 1 to 30 day exposure of rats to the hypokinetic conditions, RNA content was found to decrease on the 1st, 3rd, and 5th day and to return to the initial level by the 7th day. No changes in RNA content were observed during the subsequent stages of the xperiments. The volume of the nerve cells declined on the 3rd and 5th day, whereas RNA concentration reduced on the 1st, 3rd, 5th, and 30th day

Gorbunova, A. V.

English

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tNASA TECHNICAL MEMORANDUM
	 NASA TM-76200
RNA CONTENT IN SPINAL CORD MOTONEURONS
DURING HYPOKINESIA
A. V. Gorbunova
Translation of "Soderzhaniye RNK v motoneyronakh spinnogo mozga pri
gipokinezii", Tsitologiya, vol. 13, No..l(Jan), 1971, pp 83-87
(NASA-Th-76200) RNA CONTENT IN SPINAL CORD
NOTONEURONS DURING HYPOKINESIA (National
Aeronautics and Space Administration) 9 p
HC A02/M F A01	 CSCL 06S
G3/52
N80-290,Z6
Unclas
28199
NATIONAL AERONAUTICS AND SPACE ADMINISTRATION
WASHINGTON, D. C. 20546
	 JUNE 1980
iSTANDARD TITLE PAGE
t
1.	 Report No.
NASA TM-76200
2. Government Accession No. X	 Recipient's Catalog No.
t.	 Tula end Subtitle S.	 Report Data
RNA CONTENT.IN SPINAL CORD MOTONEURONS JITNF 1980
6.	 Perforating Orgenisetien CadeDURING HYPOKINESIA
7.	 Author(&) E.	 Perforating Organisation Report No.
A. V. Gorbunova' 10.	 work Unit Na.
9.	 Patloreting Organ44116n Name and Addraaa	 11. Contrast or Oran ► No.
SCITRAN NAS'd-3198
12. Typo of Report and Period CouoreaBox 5456
Santa Barbara, CA 93108 Translation
12.	 Sponsoring Agency Nome and Address
National Aeronautics and Space Administration ld. SponsorinsAgeneyCode
Washington,	 D.C.	 20546
15.	 Supplementary Notes
Translation of "Soderzhaniye RNK.v motoneyronakh spinnogo mozga pri
gipokinezii", Tsttologiya, vol. 	 13, Nopl(Jan),	 1971, pp 83-87
16.	 Abstract
The effect of a diminished motor activity of rats upon the RNA content in
a single isolated motoneuron of frontal horns of their spinal 'cord was
studied.	 Within a 1-30 days' exposure of rats tc 'the hypokinetic condi-
tions, RNA content was found to_	 decrease on the 1st, 3rd and 5th day and
to return to the initial level by the 7th day.-	 No changes in RNA content
were observed during the subsequent stages of the experiment. 	 The volume
of the nerve cells declined on the 3rd and 5th day, whereas RNA concen-
tration 'reduced on the lst,	 3rd, 5th and 30th day.
'	 ORIGINAL PAGE IS
OF POOR QUALITY
17.	 Key Words (Selected by Author($)) 16.	 Distrlbution Statement
Unclassified - Unlimited
19.	 Security Clessil. (of this report) 20.	 Security Clesoil. (ol this page) 21. No. of Pages 22.
Unclassified Unclassified 9
ii
NASA-110
UDC 5+7.963.3:612.8.015
RNA CONTENT IN SPINAL CORD MOTONEURONS
DURING HYPOKINESIAl
By
A. V. Gorbunova*
One should now consider it established that a reduction in the motor 	 /83**
function inevitably results in significant shifts in the metabolism of the
organism of man and animals (Katkovskiy, 19661 1967; Krupina et al., 19671
Portugalov et al., 1967; Fedorov, et al., 1968). There are data that the long
stay of an organism in a state of hypokinesia is the cause of disorders in the
nerve processes in man and animals (Gerd, 19631 Kr upina, et al., 1967). In the
literature one can also find information (Portugalov et al., 1967; Brumberg and
Pevzner, 1968) on a disruption in cellular metabolism in the nervous system
during restricted mobility. The purpose of this study was to investigate the
effect of hypokinesia on the RNA content in motoneurons of the spinal cord
anterior horns, since .this test can serve as an indirect indicator of the state
of nerve cell metabolism.
Material and Technique
Experiments were conducted on 35 mongrel albino male rats weighing 260-300 g.
The animals were placed in special, individual box cages of small size. The
tailsof the rats were attanhed by adhesive tape to a plastic dowel. This
created conditions under which even the prolonged stay of the animals in the box
cages did not result in -'.he development of chaf ings, bedsores and broken skin.
'The article contains material prepared for candidate dissertation.
Laboratory of Cytochemistry of the Institute of Medical and Biological
Problems of the USSR Ministry of Public Health.
Numbers in margin indicate pagination in original foreign text.
1
L.--
Thirty-five rats kept under vivarium conditions served as the control.
Spinal cord on the level of the intumescentia lumbalis was taken from the
experimental and control :nimals simultaneously on the 1st, 3rd, 5th, 7th, 10th,
14th and 30th day of the experiment. Fragments of Epinal cord 2-3 mm in size were
fixed in Carnua solution for 1 hour and poured into paraffin. The motoneurons
of the spinal cord anterior horns were isolated from sections 60 4 thick with the
help of a Fonbr yun micromanipulator under control of a MBI -3 microscope.
The RNA content in an individual, isolated cell was determined according to
the method of Edstrom, (1964). For this the isolated cell (fig. 1, see insert VII)*
was placed on a coverglass serving as the base of an oily chamber, and it was
exposed to triple (40 minutes each at 37 0C) effect of a ribonuclease solution
(0.4 mg/ml) in 0.2 M of ammonia-bicarbonate-acetate buffer with pH 7.6. Then the
extracts with the help of a micropipette made on a Fonbryun microforge were
completely transferred to a quartz slide. After evaporation the dried extracts
were dissolved in a microdrop of glycerin-phosphate buffer. The drops of solution
under a layer of pentadecane (Maksimovskiy, 1969) y.4ro pr^+^-qtered on a recording
ultraviolet microspectrophotometer MUF-5 by the scanning method with a-257 mµ.
The photometering was conducted with a 10 x lens and 0.07 mm probe. To calculate
the quantity of RNA contained in the drop the optic densities of the palladium
attenuators were recorded, and to determine the rate of movement of the table during
the scanning the scale of the object-micrometer was recorded with X =360 mµ. The
quantity of RNA in an individual cell was computed according to the method of Slagel
and Edstrom (1967) based on the similarity in the recording of the optic density
of the drop to a parabola. Each average amount of RNA content in the cell was
	
/84
determined from the results of analyzing 
-
'0-60 cells taken from 4-5 animals.
To calculate the RNA concentration in the cell the volume of the cellular body
was determined from a formula for the rotation ellipsoid The largest and smallest
diameters of the cell were measured with the help of a screw ocular-micrometer MOV1xl5.
Preliminary processing of the sections and isolation of the cells for size measure-
ment were carried out according to the method described by Giacobini and Holmstedt
(1958). Each average amount for the neuron volume was found from the data of
measuring 100-120 cells taken from 4-5 animals. All the numerical material was
processed by the method of var iational statistics according to Student-Fisher.
* LTranslator's notes insert not included in original.
2
For cytochemical analysis the sections of spinal cord 7 µ thick were stained
according to the method of Brashe and Eynarson.
Results and Discussion
Changes in the content and concentration of RNA were noted already on the
first day after the beginning of the experiment. Both the concentration and the
content of RNA in the nerve cells of the anterior horns of the experimental
snimals in the first five days of the experiment were lower than in the control
animals (see fig, 2, a, c,1 see table). Further, at later periods, all the way
up to the 15th day, no changes were observed in the content and concentr a%ion of
RNA in the motoneurons. On the 30th day of hypokinesia a distinct decrease in RNA
concentration in the spinal cord motoneurons was again noticed.
Observation of the animals' behavior during the experiment showed that during
hypokinesia two phases of motor activity are noted. The first phase lasts 1-5 days /85
and is characterized by the animals trying to free themselves from the unaccustomed
conditions, which was accompanied in our experiments by an increase in motor activity. 	 j
Further the animals seemingly became used tu the.new living conditions. The high
&otor activity of the animals in the period from 1 to 5 days of the experiment,
apparently, can be evaluated as the cause of the drop in RNA content and concen-
tration in the motoneurons of the spinal cord anterior horns and qualified as
"the cytochemical equivalent of high functional stress and even overexcitation of
these structures" (Portugalov at al., 1967). This is even more likely since there
is published information on the link between the functional activity of the cell
and the RNA content in it (Hyden, 19431 Hyden, 19631 Geynisman, 19651 Brodskiy,
1966) .
Further, by the 7th day of the experiment, apparently, the animals adapt to
the conditions of hypokinesia, i.e., a state occurs during which catabolism and
anabolism of RNA are balanced well with each other 1 consequently, there are no
changes observed in the RNA content and concentration. Staying under conditions
of hypokinesia results in a decrease in tl1e RNA concentration in the nerve cells
by the 30th day of the experiment, although its content remains unchanged. The
mechanisms for these phenomena have still not been disclosed.
3
r	 ^^	 °	 ?.
i	 90
60
70	 J/
f0
'	 50
t	 3	 S	 7	 fll	 f4	 ?!I
b
-	
e
90	 IN
SO . Y
70
60
—L—
SO 1 3 s 7
	 f0	 14	
— — -	
30
-	 BC
TO	 _	 -
so
AO-^
70
60
50 -
f	 3 .S	 7	 10	 fl,	 30
Figure 2. Change in Content, Concentration of RNA and Volume of Motoneuron
of Anterior Horns of Spinal Cord in Rats at Different Periods of
Hypokinesis
On x-axis--periods of h kinesia (in days); on y-axist a--RNA content (in
b--volume of cell (in % I c--RNA concentration (in %)t 1--experiment; 2--control
Parallel to a determination of the RNA content in the cell measurements
were made of the volume of the motoneurons. On the first day of the experiment
no reliable changes in the volume of neurons were obtained. On the third day
of restricted mobility a distinct reduction was noted in the volume of the
motoneur ons that occurred up to the fifth day inclusively (fig. 2, bt see table).
In the subsequent hypokinesia in the periods from 1 to 4 w--3ks (7, 10, 14, 30
days) there were no reliable shifts in the volume change (fig. 2, bt see table).
According to the data of a number of researchers (Borovskaya, 19351 Hyden,
1963 1
 Zaguskin, 1964; Geynisman, 1965; Brodskiy, 19661 Khaydarliu, 1967) there
is a clear correlation between the volume of the nerve cell body and its funetirrnal
condition. However, the evaluation of the nature of changes in the volume of
4
^i
i
i
a
i
CONTENT AND CONCENTRIMON OF RNA IN MOTONEURONS OF SPINAL CORD ANTERIOR HORNS
OF RATS IN DIFFERENT PERIODS OF HYPOKINESIA
Periods of Variant	 RNA content in Volume of cell Tit1 RNA concentration
hypokinesi	 cell (in µµg) 43 x 10-3)	 I in cell (in %)(in days)	
^y	----- - - - -
Xper lmen	 11 4 r,.7	 °U.s 0.753	 if -^ 0.1s
Ontr Ol
	
11 ± 77 (o.o_ <^ <o.a > :o.. t 0.11'1	 t> o.n:.)	 67	 o.e : 	 (o.nf)
3	 xper iment J0 31.1	 10.1 i 0.4cs	 "`,GS 0.040
(Control	 I (,... -4 33.t	 (o.00.)	 :o. * 1.57 (an^<r<o.^)	 n.nMMS 	(^.o'f)5	 ,E xper iment	 4--'I ± 24.4	 14.1 t 0.61	 o ;.;1
LControl	 u.i t3.1	 (n.0)	 _„	 (0.04	 (().()2P <o.ns
flE^ 	 t	 t O.hi^..5 0.U,^.37	 xperiment [aiyt 1 . i:	 1	 f
 
7.1 * 0.213	 H j 0.317
^COritrO1	 :,(:, 1 '.3	 ( 0.0'.)	 1	 t 1f.7; t	 (> U.O;)	 ^.+ 1 t O.03 	( 0.04
10 	per invent	 4.;, , 4.21
	 11 4- 0.3kl1	 <.:^
 0.k t;Control	 113 } S J :	 (> 0 ' U' ' )	 1' .	 ('.91 i	 (> 0. 	 11.1) .
4	
_ 	 ^
1	 xperiment i : *,7 t 111.1	 1 :L2 ,,_ 	 <.t n.nJ
gontrol	 4W. f 7.9,;	
(^U.U;.'
	 21.7_Z ^.1J	
(".1,'u',)
	
_.u,, u•nf:
	
(, U.'.•)
30	 ^xper tment j	 32,.7	
a._ 
t o.c; c	 I %^
Note: The value of D is given in parentheses.
nerve tissue cells with a change in the functional state of the organism is
still the object of discussion. Thus, some authors assume that no functionally-
governed changes exist in the dimensions of the cell body (Hyden, 1943). The
researchers who believe that this link does exist, cannot come to a unified opinion /86
on its nature. Some of them believe that the increase in the functional
activity of the cell is invariably accompanied by an increase in its volume
(Edstrom, 1957); others, on the contrary, believe that a reductions in volume
accompanies an increase in function (Borovskaya, 1935). There is also the
opinion that the increase in functional activity results in an increase in the
volume of the cellular body, which is replaced by a reduction in volume as the
cell becomes fatigued (Brodskiy, 1966; Geynisman, 1966). In our experiments,
in the period of high motor activity of the animals, apparently, fatigue of the
neurons occurred which, in all probability, also was the cause of a decrease in
volume of the cellular bodies.
The reduction in volume of motoneurons on the 3rd and 5th day of restricted
mobility can evidently be explained by the change in the distribution of water
and ions, possibly associated with the change in the physicocl.emical properties
of the membranes in the cellular structures (Brattgard at al., 1957) or change
in the content of macromolecules, in the first place, cellular proteins (Ryden,
1943).
5
Changes in the RNA content in the calls can also be established with the
use of cytological equipment (methods of Eynarson and Brashe). It follows from
figure 3 (see insert VII)* that the stay of the animals under conditions of
hypokinesia induces a reduction in the RNA content as compared to the control
level in the motoneurons of the spinal cord on the 1st, 3rd and 5th days of the
experiment.
Conclusions
1. When rats are kept under conditions of hypokinesia from 1 to 30 days a drop
is established in the RNA content in the motoneurons of the spinal cord anterior
horns on the 1st, 3rd and 5th day of the experiment, and its return to the initial
level by the 7th day of the experiment. In the further periods of the experiment
no changes were found in the RNA content.
2. A reduction in RNA concentration was found in the motoneurons of the spinal
cord anterior horns on the 1st, 3rd, 5th and 30th days of the experiment.
3. On the 3rd-5th day of hypokinesia a reduction occurred in the volume of the
nerve cells.
References
Borovskaya, A. Ya., "Functional Shifts in the Morpnology of the Neuron," Byu11.
Vsesoyuzn, inst. eksper. mad., 4 (1935), 11-14,
Brodskiy, V. Ya. Trofika kletki ["Cellular Trophism"], Moscow, 1966.
Brumberg, V. A.1 and Pevzner, L. Z. "RNA Content in Neurons and Neuroglia under
Conditions of Hypodynamics," Tsitologiya. 10, 11 (1968), 1452-1457.
Geynisman, Yu. Ya. "Shifts in Content and Concentration of Cytoplasmic Ribo-
nucleoproteins during Functional Changes in Neurons," Arkh. patol., 27, 10
(1965), 76-78.
Geynisman, Yu. Ya. "Data for Analysis of Functionally-Determined Chan g es in
Dimensions of Body and Nucleus of Nerve Cells," Tsitologiya, 8, 3 (1966),
348-358.
Gerd, M. A. "Data on Behavior and Certain Functions of Organism of People under
Conditions of Restricted Mobility," in Avl-^tsionna a i_ kosmicheskaya_
meditsina ["Aviation and Space Medicine" , Moscow, 1963, P. 126-131.
Z aguskin, S. L. "Functional Dynamics of Nissl Matter in Receptor Neuron of
River Crayfish," Tsitolo&^y, 6,6 (1964), 741-743.
* [Translator's notes insert not included in original.]
6
Katkovskiy, B. S. "Effect of Hypokinesia on Oxygen Pattern of Man__during Physical
Work," in Kislorodnyy rezhim or an	 PgQ reaulirovaniv Le "Oxygen Pattern
of Organism and Its Regulation" , 1966, 285-290.
Katkovskiy, B. S. "Main Metabolism during Prolonged Restricted Motor Activity
in Man," Kosm. biol. mad., 1, 5 (1967), 67-71.
Krupina, T. N.I Tizul, A. Ya.I Boglevskaya, N. M.I at al., "Change in the
Function of the Nervous System and Certain Analyzers in the Complex Effect
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61-65.
Maksimovskiy, L. F. "Technique for Determining Ultramicroquantities of RNA in
isolated Cells," Tsitologlya, 11, 4 (1969), 522-530.
Portugalov, V. V.f Gazenko, 0. G.; I1'ina-Kakuyeva, Ye. I.; at al., "Certain Effects
Occurring during Hypokinesia," Kosm. biol. mad., 1, 6 (1967), 18-24.
Fedorov, I. V.1 Milov, Yu. I.; V inogr adov, V. N.{ and Grishanina, L. A. "Body
Weight and Synthesis of Proteins in Animals during Hypokinesia," Kosm. biol.
mad., 2, 1 (1968), 22-24.
Khaydarliu, S. Kh. "Change in Volume of Body of Sensory and Motor Neurons in
Different States of the Nervous System," Tsitologiya, 9, 6 (1967), 644-652.
Hyden, H. "Neuron," in Funktsional'na a morfologi_ya_ uetki L"Functional Cellular
Morphology"], Moscow, 19 3, 185-248.
Brattgard, S.1 Edstrom, J. E.1 and Hyden, H. "Chemical and Structural Changes in
Nerve Regeneration," in Metabolism of the Nervous System T issues
 London,
1957, 425-428.	 —	 — —
Edstrom, J. E. "Effects of Increased Motor Activity on the D2mensions and
Staining Properties of Neuron Soma," J. Comp. Neurol., 107 (1957), 295-304.
Edstrom, J. E. "Microextraction and Microelectrophoresis for Determination and
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Physiology,_ London, 1964, 417-447.
Giacobini, El and Holmstedt, B. "Cholinesterase Content of Spinal Cord," Acta.
Ph ssiol. Scand. , 42 (1958) , 1-12.
Hyden, H. "Protein Metabolism in the Nerve Cell during Growth and Function,"
Acta Ph_ ysiol. Scand, 6, suppl. (1943), 17-23.
Slagel, D{ and Edstrom, J. E. "A Rapid Method for Calculation of the Total Amount
of Absorbing Substance in Microextracts," J. Cell. Biol., 34 (1967), 395-398.
7


RNA content in spinal cord motoneurons during hypokinesia

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