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Structure and function of the snail statocyst system after orbital missions on foton M-2 and M-3

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  • Institute of Higher Nervous Activity and Neurophysiology RAS

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In terrestrial gastropod snail Helix lucorum L. we studied the changes after a 16-day (Foton M-2) and 12- day (Foton M-3) exposure to microgravity in: behavior, neural responses to adequate motion stimulation, intersensory interactions between the photo- and the statocyst receptors, and in expression of the HPeP and FMRFa genes in the statoreceptors. Experiments were performed in the interval 13-30 hours after landing. In behavioral experiments it was found that the latency of body position response to sudden orientation change (90° pitch head-down from horizontal position) was significantly reduced in the postflight snails. Responses recorded extracellularly from the statocyst nerve to adequate motion stimulation in the postflight snails were independent of the motion direction, while in the control animals differences in responses to different directions were observed. In electrophysiological recordings it was possible to distinguish firing patterns of up to 11 of the 13 receptors that constitute the statocyst. A significantly higher firing rate in statocyst responses to body orientation at all tested speeds were observed in postflight snails, while in control snails similar dependence of statocyst responses on speed of body position change was observed, but firing rate at each speed was significantly less. Significant differences in the HPeP gene mRNA expression pattern in the statocyst receptor neurons were observed between postflight and control snails. No differences in expression of FMRFa gene expression was noted in the nervous system or statocyst after the flight. Results suggest a possibility to describe the subcellular mechanisms of changes in gravireceptors due to microgravity exposure using this simple model animal.
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STRUCTURE AND FUNCTION OF THE SNAIL STATOCYST SYSTEM AFTER
ORBITAL MISSIONS ON FOTON M-2 AND M-3
P.M. Balaban(1), A.Y. Malyshev(1), V.N. Ierusalimsky(1), N.A. Aseev(1), T.A. Korshunova(1), N.I. Bravarenko(1),
M.S. Lemak(1), M.V. Roschin(1), I. S. Zakharov(2), Y. Popova(3), R. Boyle(3)
(1)Institute of Higher Nervous Activity and Neurophysiology, Russian Academy of Sciences, Butlerova 5A, 117485,
Moscow, Russia, balaban@ihna.msk.ru; (2)Koltzov Institute of Developmental Biology RAS. ul. Vavilova 26, Moscow,
119991 Russia; (3)NASA BioVIS Center, Ames Research Center, USA, richard.boyle@nasa.gov
ABSTRACT
In terrestrial gastropod snail Helix lucorum L. we
studied the changes after a 16-day (Foton M-2) and 12-
day (Foton M-3) exposure to microgravity in: behavior,
neural responses to adequate motion stimulation,
intersensory interactions between the photo- and the
statocyst receptors, and in expression of the HPeP and
FMRFa genes in the statoreceptors. Experiments were
performed in the interval 13-30 hours after landing. In
behavioral experiments it was found that the latency of
body position response to sudden orientation change
(90° pitch head-down from horizontal position) was
significantly reduced in the postflight snails. Responses
recorded extracellularly from the statocyst nerve to
adequate motion stimulation in the postflight snails were
independent of the motion direction, while in the control
animals differences in responses to different directions
were observed. In electrophysiological recordings it was
possible to distinguish firing patterns of up to 11 of the
13 receptors that constitute the statocyst. A significantly
higher firing rate in statocyst responses to body
orientation at all tested speeds were observed in
postflight snails, while in control snails similar
dependence of statocyst responses on speed of body
position change was observed, but firing rate at each
speed was significantly less. Significant differences in
the HPeP gene mRNA expression pattern in the
statocyst receptor neurons were observed between
postflight and control snails. No differences in
expression of FMRFa gene expression was noted in the
nervous system or statocyst after the flight. Results
suggest a possibility to describe the subcellular
mechanisms of changes in gravireceptors due to
microgravity exposure using this simple model animal.
1. INTRODUCTION
Following space flight of 1-2 weeks many
astronauts/cosmonauts experience conditions that can be
attributed to gravireceptor dysfunction, e.g., illusionary
feelings, vertigo, nausea and vomiting, gaze fixation
disorder and ocular nystagmus. Microgravity (µG)
exposure results in changes in both the linear and
angular vestibulo-ocular reflexes, and the spatial
orientation of eye velocity towards gravity requires days
to recover [4]. Typically within days to weeks postflight
these sensations recede, indicating a neural re-
adaptation to 1G has occurred. Understanding the
cellular mechanisms underlying the adaptive and re-
adaptive responses of the equilibrium organ is essential
in the design of specific countermeasures to mitigate
adverse reactions to the space environment. Our
hypothesis is that spaceflight challenges the gravi-
receptors and causes changes in their cellular function,
that include changes in the regulation of specific gene
expression and in the electrical behavior of the
receptors, and that re-adaptation to 1G is a time process
that can be tracked by intracellular measurements.
In this study we used the terrestial snail Helix lucorum
Linnaeus (Pulmonata, Gastropoda). These snails are
small, strong and can remain metabolically active under
restrained conditions for weeks. Snails have flown on
shuttle, MIR and ISS missions in the past. Behavior and
cellular networks underlying it are well described in this
animal [1].
In order to reach the goal, we investigated the statocyst
receptor function as a consequence of µG, in particular
the re-adaptation phases in 1G, using different
approaches such as analysis of changes in gravity-
dependent behavior, expression of a specific gene
associated with the statocyst, and electrophysiological
analysis of changes in statocyst responses to
gravitational stimuli. Report on Foton-M2 flight was
published [2], and in the present study we describe new
data from the Foton-M3 flight.
2. METHODS.
Twenty small (1-3 gm), Helix aspersa, and 16 large (3-8
gm) snails, Helix lucorum, were flown on unmanned
Russian satellite Foton-M3. From the postflight
conditions of the enclosure it was apparent the snails
were active during most, if not the entire, the mission.
Temperature profiles provided indicated that the snails
were exposed to an optimal temperature (16-20oC). The
juvenile snails were fixed for pedal peptide analysis
using mRNA expression analysis upon receipt of the
snails at 14h after landing. After weighing the adult
snails, behavioral tests (14-18 hrs after landing) were
conducted to measure their timed negative gravitaxis
response to tilt. After the behavioral tests, the snails
were used in an electrophysiological study (15-20 hrs
after landing) to evaluate the intersensory interaction
between the photo- and statoreceptors using natural light
and a tilt stimulus at different speeds in the isolated
snail central nervous system (CNS) preparation, and an
electrophysiology study to evaluate the firing rate
modulation of the statoreceptor to tilt. Conventional
electrophysiological techniques were used. Synchronous
ground control snails were exposed to the same
_________________________________________
Proc. of the 'Life in Space for Life on Earth Symposium', Angers, France
22–27 June 2008 (ESA SP-663, December 2008)
Fig. 1. Results of behavioral testing of control and
postflight snails in “negative gravitaxis” experimental
situation. Inset shows phases of the snail stereotypic
response to sudden shift of the snail with platform from
horizontal to “head down” position. The plot shows
averaged (±SEM) time of the behavioral responses at the 2
phases for the negative gravitaxis response for the 5 flight
and 6 control snails. Flight snails were significantly faster
in their response to pitch stimulation at each phase.
temperature profile as the flight snails. Cooling of
animals to 4°C and injection of isotonic MgCl2 were
made before the CNS isolation to minimize pain. All
efforts were made to minimize quantity of used animals.
Experimental protocol was in accordance with the USA
NIH “Guide for the Care and Use of Laboratory
Animals”.
3. RESULTS
3.1. Behavioral experiments.
The behavioral "negative gravitaxis" responses of the
snails showed that the flight snails in general responded
faster than their control counterparts. Inset on Fig. 1
shows the test procedure: phases 1 and 2 (T1 and T2)
represent the epochs of behavioral response (fast re-
orientation to the normal for the snail position – “head
up”) following a horizontal to vertical head-down pitch
of the snail. Comparison (Fig. 1) of averaged latencies
of the behavioral responses at the two phases for the
negative gravitaxis response for the 5 flight and 6
control snails showed significant differences in
performance. Flight snails were faster in their response
to pitch stimulation, and this was significant (p <0.04;
nonparametric, two-tailed, unpaired Mann-Whitney
test). Results clearly suggest the existence of changes in
behavior of the postflight snails and completely coincide
with results of Foton-M2 mission [2].
3.2. Postflight changes in gene expression in CNS
3.2.1. In situ hybridization with probes to the gene
encoding FMRFamide.
Expression pattern of the preproFMRFa gene was
investigated in the postflight and control snails using
specific mRNA expression techniques. It was found in
accordance with published data that FMRFa is
expressed in all circumesophageal ganglia in control and
postflight nervous systems. No expression was seen in
statocyst organs. Pattern of expression was very stable
and similar in control (8 snails) and postflight (8 snails)
preparations. Thus, participation of this small
neuropeptide in vestibular changes was not confirmed.
3.2.2. In situ hybridization with probes to the Pedal
Peptide snail gene.
Expression pattern of the preproHPep gene [7] was
investigated in the postflight and control snails using
mRNA expression techniques. The pedal peptide is
expressed in the primary statocyst receptor cells under
physiological load [7], and regulation of this gene
expression pattern might signal how the statocyst
receptor is "tuned" by the gravity vector. Specifically
stained cells (expressing the HPep gene) were observed
in cerebral, subesophageal ganglia complex and in pedal
ganglia in all preparations. No systematic differences
were observed in postflight snails relative to the control
snails in location and pattern of the stained ganglion
neurons (data not shown). On the contrary, a qualitative
difference in staining of the statocyst neurons was
observed. In the control animals (11 snails, 22 statocysts
examined) the 3 neurons expressing preproHPep gene
were found in 59% of cases, while in 16 postflight snails
fixed 12 hours after the landing, expression was found
in 96% of cases. There are only 13 neurons in total in
the statocyst, and up-regulation of gene expression in
several of them represents possible changes in their
function. This specific increase in gene expression in
statocysts is indicative of the physiological load and
may reflect the flight experience. These results fully
coincide with results from Foton-M2 flight [2].
3.3. Electrophysiological experiments in isolated
CNS.
Analysis of the background activity (spontaneous firing)
in the statocyst nerve was performed similarly in control
and postflight snails. For each preparation three 10s
intervals from recordings under conditions “light on”
(switched on 50s before the analyzed interval) and in
darkness were taken for analysis. Vestibular stimulation
was not presented for at least several minutes before the
first analyzed interval. Data from 6 flight and 6 control
snails were analyzed. No significant difference in level
of background activity was observed in darkness, and in
the condition “light on”. In the snails from the Foton-
M2 flight the firing rate under conditions “light on” was
significantly different. The nature of the conflicting
results in this test is unclear, but may reflect some basic
difference in the snail populations and/or conditions.
Analysis of the responses to different directions of
Fig. 3. Electrophysiological responses to motion in
statocyst nerve in isolated CNS of 6 control and 6
postflight snails. Upper plot shows responses in control
and postflight snails to rise of the platform (start at zero
point) with “head down” orientation. Duration of rise is
shown. Significant difference was observed (p<0.01,
pairwise comparison, Mann-Whitney).
motion was performed using the same isolated CNS
preparations. The platform with isolated CNS and
recording chambers was tilted to 19° up from horizontal
position and back (Fig. 2). Time of rise and fall varied
from 550ms to 3050ms.
Orientation of preparation was identical in all
experiments and the rise of platform corresponded to a
head down tilt in intact snails, as was made in
behavioral experiments. Three consecutive rises and
falls were made at 30s intervals in darkness, followed
by a similar procedure was performed with light on. The
preparation was then rotated 180° and the protocol was
repeated. Averaged extracellularly recorded statocyst
nerve firing rates in the 6 postflight and 6 control snails
were analyzed to rise and fall steps. Responses were
quantified by calculating the number of spikes per 0.3s
bin (Fig. 3, upper plot). It is evident from the results that
in postflight animals the rate of spike discharge is
significantly higher at different speeds (Fig.3), nearly 3-
fold at the fastest test speed (upper graph) and nearly 2-
fold at the slowest test speed (lower graph). A similar,
but not significant, tendency was observed after Foton-
M2 flight [2].
Comparison of different direction of tilt showed that in
control animals the response to rise was significantly
and constantly higher than to the fall of the platform. It
reflects an orientation selectivity of the animal that
normally exists and is the basis of negative gravitaxis.
Similar responses in postflight animals were not
constantly selective. Responses to rise at first are
smaller, then became bigger (Fig. 4). In order to
illustrate this effect, we plotted a difference between the
responses in two orientations of the platform. Responses
from each preparation to the rise and fall of the platform
were subtracted. Thus, if no preference to orientation
exists, the response will be close to zero. It is evident
from the results that there exists an orientation
selectivity of responses to tilt (difference is significantly
away from zero) in both groups of snails, and a
significant difference between flight and control snail
(statistical significance was evaluated by RM-ANOVA
with post-hoc analysis, p<0.01 for several time points)
exists also. Orientation selectivity in controls (more
spikes in response) corresponded to head down position
of the snail, while in postflight snails the selectivity was
not so clear because within the first bins the response
favored head up, then the response changed to head
down. It suggests that the neural responses of the
statocyst nerve to adequate motion stimulation in the
postflight snails is independent of the motion direction,
while in the control animals significant differences in
responses to different directions were observed. Similar
results were observed after Foton-M2 flight [2].
4. DISCUSSION.
In this study we investigated the function of the gravi-
sensing statocyst receptor during the re-adaptation phase
after landing from a 12-day orbital mission. A
behavioral motor response called negative gravitaxis
revealed a more rapid display of this behavior in
Fig. 2. Scheme of experimental set-up for recording
vestibular nerve activity.
Fig. 4. Electrophysiological responses to motion in
statocyst nerve in isolated CNS of 6 control and 6
postflight snails. The plot gives the averaged difference
between statocyst nerve responses to rise and fall stimuli
correspondint to “head-up” and “head down” positions,
see inset. A response near zero indicates no preference. In
control snails a preferred direction of tilt exists, while
selectivity is not similar (changes direction) in the
postflight snails. A significant difference between flight
and control snail was observed in the middle of the tilt
(p<0.01, RM-ANOVA with post-hoc analysis).
postflight than in control snails. A qualitative
upregulation in the expression of a peptide associated
with the statocyst was seen. Lloyd [5] first identified
this pedal peptide in sea slug Aplysia californica.
Although its role in the nervous system remains
unknown, the beat frequency of cilia in epithelial cells
of Tritonia is modulated by this peptide [8]. In 1997 the
gene encoding pedal peptide was identified in terrestrial
snail Helix lucorum [7]. According to in-situ
hybridization data this gene is widely expressed in the
nervous system of Helix and as well in some
statoreceptor hair cells. Taking into account the known
effect of pedal peptide on cilia beating and muscle
contraction we can assume that it might play an
important role in the regulation of mechanical
transduction processes of the hair bundles of
statoreceptors, such as regulation of actin-myosin
complex within the cilia, and the observed change in
expression of gene encoding this peptide suggests
existence of lasting influence of the flight.
Another feasible interpretation of HPep data is the
known mosaic expression of different peptides in the
statocyst hair cells [6] that is believed to code different
position or movement information. In this case the HPep
expression from an almost negligible level to a
significant level may indicate a change in encoded
information transmitted to the nervous system from
vestibular organ, and reflects the behavioural changes
observed in postflight snails.
Physiological response in the gravito-inertial sensing
organs can occur rapidly in an organism in direct
response to a change in gravitational force: a
hypersensitivity to translation acceleration was seen in
fish otolith afferents within the first day following
orbital missions [3]. The results here in the isolated
statocyst in invertebrate (Fig. 3) are directly in line with
the vertebrate data, and conform to the proposition that
µG exposure leads to changes in gravireceptor function
(Figs. 3 and 4). Disappearance of orientation selectivity
of statocyst neurons in postflight snails evidently
reflects an overall change elicited by microgravity.
In summary, following a 12-day orbital unmanned
mission, significant changes were observed in the
snail’s behavior, expression of a gene encoding a pedal
peptide, orientation selectivity of statocyst responses,
and an upregulation of the response sensitivity to
motion. We assume that the changes were the result of
µG exposure. Thus, the results obtained in this simple
animal model open the possibility for identifying other
neurobehavioral responses and even revealing
subcellular processes affected by the space environment.
5. REFERENCES
1. Balaban P.M. Cellular mechanisms of behavioral plasticity
in terrestrial snail. Neurosci Biobehav Rev. Vol. 26, 597-
630, 2002.
2. Balaban PM, Malyshev AY, Zakharov IS, Aseev NA,
Bravarenko NI, Ierusalimsky VN, Samarova AI, Vorontzov
DD, Popova Y and Boyle R (2006) Structure and function
of the snail statocyst system after a 16-day flight on Foton
M-2. J Grav. Physiol.13: P201-P204.
3. Boyle R., et al. Neural readaptation to 1G following return
from space. J. Neurophysiol., Vol. 86, 2118-2122, 2001.
4. Cohen B., et al. Vestibular experiments in space. In,
Experimentation with animal models in space. Edited by G.
Sonnenfeld. Elsevier B.V., pp. 105-164, 2005.
5. Lloyd P.E. & Connolly C.M. Sequence of pedal peptide: a
novel neuropeptide from the central nervous system of
Aplysia. J Neurosci. Vol. 1, 312-317, 1989
6. Ohsuga K., Kurokawa M., & Kuwasawa K. Mosaic
arrangement of SCPb-, FMRFamide-, and histamine-like
immunoreactive sensory hair cells in the statocyst of the
gastropod mollusc Pleurobranchaea japonica. Cell Tissue
Res. Vol. 300, 165-172, 2000.
7. Poteryaev D.A., et al. Characterization of cDNA clone
encoding pedal peptide in terrestrial snail. Neuroreport.
Vol. 8, 16, P. 3631-3635, 1997.
8. Willows A.O.D., et al. Modulation of ciliary beat frequency
by neuropeptides from identified molluscan neurons. J Exp
Biol. Vol. 200, 1433-1439, 1997.
... Up-and down-regulation of peptide expression in the CNS in response to external stimuli (including injury) has been poorly studied so far and seems not to be a typical feature for most of the neuropeptides. Previously, we have shown that a specific factor (microgravity) can selectively activate pedal peptide synthesis in the cells of snail's statocyst, an organ responding to gravitational stimuli, while no changes were detected in the CNS neurons containing the same neuropeptide (Balaban et al. 2006). In the silkworm, Hagino et al. (2010) have shown a close correlation between the immunoreactive intensity of FXPRLa peptides and environmental conditions. ...
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Experimentation with animal models in space
  • B Cohen
Cohen B., et al. Vestibular experiments in space. In, Experimentation with animal models in space. Edited by G. Sonnenfeld. Elsevier B.V., pp. 105-164, 2005.