ABSTRACT
The ability of limpets to exhibit high tenacity
in clamping down on rocks for prolonged periods of time
is accounted for by a possible catch mechanism. Contractile
properties of a preparation of the shell-foot retractor
muscle of the limpet Lottia gigantea were investigated
using salines of altered ionic concentrations and known
muscle effectors (caffeine, acetylcholine, and 5-hydroxy-
tryptamine). The following results were obtained;
1. The shell-foot retractor muscle exhibits twitch responses
and summation characteristic of most muscle types.
2. Acetylcholine, caffeine, high K'saline and high K-
caffeine saline induce muscle stress in the absence
of electrical stimulation.
3. 5-HT reduces muscle stress in the absence of electrical
stimulation.
4. High Ca, low Na, caffeine, high K, high K-caffeine,
a hypertonic solution, Ach and 5-HT potentiate a
contraction in response to electrical stimulation.
The responses vary between test salines and depend
on prior response to electrical stimulation and
exposure time of 'the muscle to the test solution.
5.
Acetylcholine alone induces a sustained contraction
stress. 5-HT relaxes this contraction at a relaxation
rate 33 times the rate in acetylcholine alone. This
pharmacologic response is evident in muscles known
to exhibit a catch mechanism of contraction.
A maximum contraction stress of 3.1x 10° N/m was
measured in the shell-foot retractor muscle. To obtain
an indication of the muscle contribution to limpet
tenacity this value was compared to the stress necessary
to dislodge a tightly clamped limpet from an intertidal
rock and found to be of equal magnitude.
INTRODUCTION
Molluscan muscle has the capability to develop great
tensions and sustain prolonged contraction stresses
through a "catch" mechanism. This mechanism has been
extensively studied in the anterior byssus retractor muscle
(ABRM) of Mytilus edulis (Twarog 1954,1960,1967,1967a, 1968.
1969,1971,1972) and it is believed to be a type of mechanism
available to many muscles which may develop specialization
for catch to varying degrees (Wilson and Larimer 1968). I
examined the possibility that a catch mechanism exists in
limpet shell-foot retractor muscle. The existence of such a
mechanism might account for limpets' ability to remain
tightly clamped down on a rock for extended periods of
time when faced with wave shock or dessication stresses.
This work is the first to study the properties of limpet
muscle. I began by examining the muscle response to single
and repeated electrical stimulation of varying strength,
duration, and frequency. Interest in the catch hypothesis
then led me to examine the pharmacological properties of
the muscle. By altering a standard artificial seawater
solution, the effects of relative concentrations of major
ions were tested on the muscle, using the solution alone
and accompanied by electrical stimulation; changes in the
contraction stress and relaxation times were noted, Known
muscle effectors such as caffeine, acetylcholine and 5-HT
salines were also used to examine contractile properties
of the muscle preparation. Data suggestive of catch is
discussed as are
effects
of certain test salines on the muscle contraction stress.
MATERIALS AND METHODS
Lottia gigantea were collected at Pacific Grove, Ca..
and kept at 14 degrees Celsius in circulating seawater.
Limpets were injected with 5 ml MgCl, solution (1 part
I M MgCl, to 2 part seawater) to induce relaxation in the
foot muscle. With the limpet on its dorsal side, longitudinal
cuts 5mm apart were made on the shell-foot retractor muscle
from its point of attachment on the shell to the ventral
surface of the foot (10-12 mm depending on the size of
the limpet). The muscle segment was then teased away from
the shell and the most lateral layer (lmm) was used as the
final preparation (dimensions: 10mmx 5mmx lmm). The
muscle preparation was placed in running seawater for 1
hour to allow recovery from the anesthetic.
The preparation was mounted horizontally between two
flattened alligator clips. One clip was fixed to the bottom
of the dish containing the test solution. A force transducer
was attached via a fine chain to the free-moving clip,
The sample was stretched by approximately 202 (2mm) above
its dissected length to return it to its in vivo length
(as measured in the intact limpet prior to dissection).
This extension required a force of 0.2 N(a stress of
4 x 10" N/m*). An additional loading force of O.25N
was applied (total loading stress of 9x 10' N/m.
A Grass S44 stimulator and a bipolar electrode were used
to deliver electrical stimuli to the muscle.
A standard artificial seawater solution (ASW) was used
as a control (470 mM Nacl, 10 mM KCl, 10 mM Cacl,, 5 mM Hepes,
pH - 8.1) to test the following altered salines: High K
(400mM KCl, 80 mM Nacl), caffeine (30mM), high K-caffeine
(30 mM in high K), high Ca (replaced Mgcl with Cacl.),
low Na(replaced NaCl with Triscl), 5-hydroxytryptamine (10-4M).
acetylcholine (10 "M) and a hypertonic solution (300mM in
ASW). All solutions were titrated to pH of 8.1 with .5M Naoh
and stored at 4 degrees Celsius.
Experiment 1
Single and twin shocks of 12 Volts, 50 millisecond pulse
duration were applied to muscle preparations in ASW. The
interval between double stimuli was varied between 2 and
10 seconds.
Experiment 2
The rate of repeated stimulation was varied from 1 to 10
pulses per second at 15 Volts, 60 ms pulse duration for
33 second applications.
Experiment 3
The effect of various solutions on the stress developed by
the muscle were investigated. Each solution was tested on
8 muscle samples. Each muscle sample was used to test only
one solution and its ASW control. Solution baths were
maintained at 15 degrees Celsius and were flushed every
10 minutes (except when indicated) in the following order:
ASW, test solution, ASW, test solution. The effect of
the solution alone on the muscle was tested without elec-
trically stimulating the muscle in the bath. Eight muscle
samples were then bathed in each solution and this time the
muscle was electrically stimulated. Series of shocks of
90 Volts, 6 ms pulse duration, 10 pulses per second, and
3 second duration were applied. At the time of maximum
tension evoked by the solution alone, the first series of
shocks was applied; a second series was applied 1 minute
later, etc. In solutions in which there was no measurable
effect on the muscle stress by the solution alone, the
first series of shocks were delivered after 2 minutes of
bathing and the second series after 3 minutes, etc.
Contraction stress, contraction times (time from baseline to
maximum peak) and relaxation times (time from peak to baseline)
were calculated for each muscle response.
Experiment 4
Repeated single shocks of 60 Volts, 6 ms pulse duration
with 10 second intervals between shocks were applied to 8
muscle samples in contact with (but not submerged in) ASW.
High K-caffeine (5ml) saline was applied directly onto the
muscle while it was still in contact with ASW.
Experiment 5
Three muscle preparations were placed in contact with Ach
saline and allowed to remain there until the muscle stress
subsided to baseline. 5-HT (5ml) was applied directly to
the muscle once the maximum muscle stress evoked by the Ach
solution was reached.
(elaxation times for Ach alone and
Ach + 5-HT trials were recorded.
RESULTS
Experiment 1: A single contraction was evoked by a single
stimulus and is therefore classified as a twitch response.
Based on criteria in Schmidt-Nielsen (1979) the waveform
obtained in the response is characterized as phasic (Figure 1)
Double stimuli set apart by short intervals result in
summation and a contraction stress higher than that evoked
by a single twith is recorded.
Experiment 2: Repeated stimulation of the muscle at 15 Volts
and 60 ms pulse duration, 1 pps, and 33 second duration
evokes a tonic waveform. Summation in the tonic mode at a
rate of I pps gives a characteristic "sawtooth" appearance.
A higher rate of stimulation (10 pps) evokes a higher maximum
stress and fusion to a tetanus (Figure 2).
The responses recorded in the first two experiments are
characteristic of most muscle types (Mattison and Arvidsson
1966; Heyer et al 1973; Schmidt-Nielsen 1979).
Experiment 3: Figure 3 shows those solutions which by
themselves affected the muscle stress. Also indicated are
the additional effects of the first series of electrical
stimuli in the given solution. The hypertonic solution,
high Ca and low Na salines alone had no measurable effect
on the muscle stress and therefore are not included in Fig.3.
Figure 4 shows the effects of the first and second series
of shocks on the muscle stress in each test solution
expressed as a percentage increase above the stress measured
in Asw (12 x 10" N/m2 in the first series and 11.4 x 104 N/m
in second series of shocks).
High Caf: The high Ca saline potentiated a contraction
stress when the muscle was electrically stimulated. Stresses
evoked by the fourth series of shocks were equal to those
evoked by the second series of shocks. This finding is
contrasted with that in ASW after the fourth series of
shocks, where the stresses were half the size of those evoked
by the second series of shocks in ASW.
Low Na: Low Na saline potentiated a contraction appro-
ximately 367 above that of ASW in the first series of
shocks. However, note that the response to the second series
of shocks in the solution shows a great decrease from the
first contraction stress.
Hypertonic saline solution: After a 5-minute bath in
hypertonic saline, the muscle responded to electrical
stimulation with a contraction stress higher than that in ASW.
In contrast to this, a 1-hour bath in hypertonic saline
depressed the muscle response to electrical stimulation
to a stress 452 below that in ASW.
Caffeine: Although caffeine itself had minimal effect
on the muscle, it appears to potentiate the highest
contraction stress of all the solutions tested in response
to electrical stimulation. The response can be repeated
provided that a rest interval of 60 seconds is allowed
between trials. However, after six repetitions, the
contraction stress evoked became successively lower
(2 x 10' N/m' after six series of shocks) than would be
accounted for by fatigue (as measured in ASW (6 x 10* N/m2).
Note that these values are those obtained by the applied
stimulus and do not include the loading stress of 9 x 10* N/m?.
High K and High K-caffeine: High K and High K-caffeine
salines potentiated tonic contractions which peaked at
40 seconds and 30 seconds respectively after the initial
contact with the muscle. The muscle responsiveness to
electrical stimuli in the High Kalone decreased to zero
after the first 4 minutes in the bath. When caffeine is
present in the high K medium however, the muscle remains
responsive to electrical stimulation for the first 7 minutes
of bathing. The relaxation time after a shock in High K
alone is decreased from 27 seconds to 8 seconds when caffeine
is in the medium (Figure 5). The relaxation time of the
phasic waveform after electrical stimulation in the caffeine
saline alone is approximately 6 seconds. Note that when
both high K and caffeine are in the same medium, the rela¬
xation time is not an average of the two but is much closer
to that measured in caffeine alone. Figure 5 illustrates
typical waveforms in caffeine, high K and high K-caffeine.
Experiment 4: Figure 6 illustrates the summation effect
of repeated single shocks of 60 Volts, 6 ms pulse duration,
10 second intervals between shocks. Note that the stress
does not increase on every consecutive shock but instead
there is a tendency to maintain a constant stress (first
10 shocks). When high K-caffeine is added directly onto
the muscle, the stress increases on each subsequent shock
(shocks + 10-15) until it reaches a maximum. This stress
decreases despite continued stimulation. The addition of
high K-caffeine during this stress decrease appears to
prevent further loss of stress and even enhances the response
of the muscle to continued electrical stimulation. I
can offer no specific mechanism for this action of high K
caffeine.
Experiment 5: Although Ach (10 *) itself had minimal
effect on the muscle, it potentiated a contraction stress of
25 x 10' N/m' when the muscle was electrically stimulated
in the solution (from Experiment 3). The second series
of shocks in Ach evokes a stress equal to that evoked
by the second series in ASW (20.4 x 10' N/m2). When the
muscle was exposed to 5-HT (10 M) alone, a 1002 relaxation
of the loading stress (9 x 10' N/m2) was observed. When
electrically stimulated, muscle in 5-HT was more responsive
than the same muscle in ASW. This is reflected in the
absolute increase in stress caused by the 5-HT saline after
electrical stimulation (18 x 10' N/m) of the muscle. Compare
this with an absolute increase in stress in ASW in response
to equivalent electrical stimulation (12 x 10“ N/m2). However.
because the shock in 5-HT was applied at an already decreasing
stress of 3 x 10' N/m2 (Figure 3).
Figure 7 illustrates the sustained contraction stress
in response to Ach and the relaxing effect induced by 5-HT.
Ach potentiated the muscle to sustain a stress of lX 10' N/m'
for 420 seconds in the absence of electrical stimulation.
a relaxation rate of 23.8 N m2/sec. The application of
3-HT at the point of maximum contraction stress (possible
catch state) evoked by Ach alone causes a relaxation to
baseline in 25 seconds, a relaxation rate of 800 N m-2/sec.
approximately 33 times faster than the rate in Ach saline
alone.
DISCUSSION
The characteristic sawtooth appearance that fuses to
a tetanus at higher rates of repeated stimulation is also
observed in Cryptochiton stelleri (Harrison 1975). By
adjusting the frequency of stimulation above the minimum
for tetanus, a several fold increase in the stress is possible.
This is in contrast to most vertebrate muscle (cat eye muscle
being an exception) where variation in stimulation frequency
has little influence on force. Note that the degree to
which a given test saline alone potentiates a contraction
varied widely between solution and further differed when
an electrical stimuli was applied to the muscle in the
bath. The mechanism of potentiation of contraction is
not clearly understood but it is likely to involve many
sites of action (Schulman and Weight 1976). The data
generated in this work cannot elucidate the details of the
potentiation mechanism but does suggest some general
aspects likely to be involved in the overall mechanism
which will be discussed in different sections.
High Ca
Direct entry and diffusion of Ca through the small
diameter of the shell-foot retractor muscle cells more
than likely accounts for muscle contraction as is found
for other molluscan muscles (Huddart et al 1977, retractor
and columella muscles). In experiment 3 the ability of
the muscle to repeatedly develop a constant muscle stress
(equal to that developed after the second series of shocks)
in response to a fourth series of shocks is suggestive of
the importance of an external Ca concentration. This
response may have been possible only in the presence of
the high Ca "of the saline. Fatigue seems to have been
significantly lowered from that observed in ASW after the
fourth series of shocks. The external Ca may have shifted
the ionic balance at the cell membrane in favor of repeated
potentiation of contractions of constant stress. Since
the high Ca levels potentiated a contraction 257 higher
than in AsW after the first series of shocks(Figure 4).
it could be said that the rate of entry of Ca into the cell
is likely to be fast enough to enable potentiation in
response to this specific electrical stimulation. If the
rate is faster than the rate required to potentiate in
response to repeated stimuli, it may account for potentiation
of twitches as well (as in skeletal smooth muscle). However.
if the rate of entry is not fast enough for twitch potentiation
then the Ca" concentration may only play a role in prolonged
contractions evoked by repeated stimuli. Atwater (1974)
found evidence for the latter in the giant retractor cells
of Megabalanus psittacus.
Low Na
Note that the low Na saline I used is actually a zero-Nat
saline (Triscl replacement). Therefore, the inability of
the muscle to develop a substantial stress on the second
series of shocks in this low Na (302 decrease below the
stress developed in ASW) suggest the need to have an external
Na concentration to allow responses to electrical stimulation
beyond the initial series of shocks. Subsequent responses
may need a Na influx such as that required in vertebrate
skeletal muscle to initiate an action potential and cause a
contraction. Enough Na may have already been present in the
immediate surroundings of the muscle cells to allow a subs-
tantial response to the first series of shocks in the low
Na saline. Experiments with high Na saline and blocking
agents such as curare might elucidate the nature of the
mechanism involving Nakions (i.e. the possible existence
of a sodium pump).
Hypertonic Saline
The performance of the muscle after a 5-minute bath
in hypertonic saline compared with its performance after a
1-hour bath in hypertonic saline indicates a loss of
responsiveness to electrical stimulation possibly due to
prolonged exposures to high tonicty (Figure 4). This
response may be experienced in vivo when the limpet is
challenged with osmotic stresses and/or dessication.
After the first two series of shocks, progressive series of
stimuli for an hour thereafter however, evoked constant
stress never below 50% of the corresponding stress in ASW.
This data indicates that although the stress evoked in
hypertonic saline is far from maximum, it does not change
sharply over time after the first hour of exposure to
hypertonic media. It would then be fair to expect that
a live limpet should be able to withstand prolonged periods
of conditions which cause its muscle to be in a hypertonic
state, despite its overall depressed ability to evoke
high contraction stress. Its tenacity might be expected
to be lower due to a depressed muscle performance; however.
the rate at which its tenacity decreases will likely remain
relatively constant over time.
Caffeine
Caffeine has been reported to modify the properties
of sarcotubular preparations (Weber and Herz 1968).
surface membrane of muscle (Bianchi 1961) and nerve terminals
(Hofmann 1969). The common denominator for these different
manifestations is an effect of caffeine that induces
changes in the distribution of Ca and modifies its movement
across cellular membrane components. Mechanisms evident
in muscles that respond to caffeine in the same way that
the shell-foot retractor muscle does suggest a Ca mobi-
lization role for caffeine. It is credited with increasing
the cell membrane permeability to Ca'tinflux and efflux.
(Sandow 1965, frog semitendinosus; Muneoka and Mizonishi
1969, Nytilus edulis; Chiarandini 1970, crayfish; Ashley
1977. Blanus nubilus). The results obtained in experiment
3 showing the successive decrease in stress after the
sixth series of shocks (a stress reduction lower than can
be accounted for by fatigue) in the caffeine saline may
suggest that the source from which caffeine releases Ca'
to induce contraction is dependent at least in part on
the external Ca concentration. A similar result in
response to caffeine was found in crayfish muscle (Chiarandini
1970). It would be interesting to treat the SFR muscle
with high Ca'and low Ca2 salines to test for any differences
in repeated response to electrical stimulation.
High Kand High K-Caffeine
The unresponsiveness of the muscle in the High Kafter
4 minutes of bathing and electrical stimulation may be an
indication that the high K increasingly depolarizes more of
the muscle cell membranes over time, thus rendering it
virtually impossible to evoke a further mechanical response
when the muscle is shocked. The shortened relaxation time
of a contraction in high K-caffeine (Figure 5) seems to
suggest that the depolarizing action of high kon the cell
membrane is either slowed down or somewhat counteracted
by the action of caffeine on the muscle. Axelsson and Thesleff
(1958) found the same result in vertebrate striated muscle.
A mechanism for this effect cannot be derived from the data
I obtained.
Acetylcholine and 5-HT
Potentiation of contractions by Ach in response to
electrical stimuli and the relaxation caused by 5-HT alone
on the shell-foot retractor muscle have also been found in
gastropod radular retractor muscles (Mattison and Arvidsson 1966).
The enhanced response to electrical stimulation in 5-HT
(concentrations below 10 6M) were also found in other gastropods
(Kobayashi and Muneoka 1980) and in an opistobranch (Cohen
et al 1978). The increase in relaxation rate caused by
S-Hr on Ach-induced stress is suggestive of a catch mechanism.
as is found in the ABRM of Mytilus edulis. Ach injections
(10 M) in the isolated ABRM initiate catch (Cole and Twarog
1971) and 5-HT(10"M), the probable transmitter substance
feleased by relaxing nerves in Mytilus (Twarog 1954,1968)
release catch. Certain features of its mode of action are
known (Cole and Twarog 1971) while other remain unclear.
Some evidence points to its effect in increasing membrane
conductance and triggering reaction cascaded to reduce the
level of free intracellular Ca tTwarog 1966), an effect
opposite that of caffeine. It may be worthwhile to conduct
an experiment to test the effect of 5-HT applications on a
muscle previously bathed in caffeine.
Table 1(adapted from Schmidt-Nielsen 1979) gives maximum
contraction stress values measured in various animals. I
measured a value of 3.1 x 102 N/m2 for the limpet shell-
foot retractor muscle. This stress was reproducible as it
did not irreversibly damage the muscle preparation. To
get an indication of the contribution the muscle contraction
stress makes to a limpet's tenacity, this value can be
compared to the stress necessary to pry a limpet off a
rock. The latter is a value of 1-4 x 10° N/m? (Denny 1985
pers.comm) and it is created by a low pressure in the mucus
of the foot when the retractor muscle contracts, thus
adhering the limpet to the rock and accounting for its
high tenacity. My value may be an underestimate due to the
nature of the muscle preparation in which it was measured.
I did not work with individual muscle fibers; therefore, the
number of fibers contracting in any one particular direction
upon stimulation, is likely a function of their angle of
orientation relative to the direction in which the contraction
stress is measured. Nevertheless, if one assumes that the
limpet is exerting maximum contraction stress when it
detects a stimulus on its shell to avoid being dislodged,
then this value of 3.1x 10° N/m reflects the ability of the
SFR muscle to account for most of the stress necessary to
keep the limpet clamped down on the rock. Factors such as
osmotic stress and/or dessication may also play an interactive
role in affecting the limpet's tenacity (Boggs and Marzuola,
pers. comm).
LITERATURE CITED
Ashley, C., Ellory, J.C., and Griffiths, P.J. (1977). Caffeine
and the contractility of single muscle fibres from
the barnacle Balanus nubilus. J. Physiol., 269,421-439.
Atwater, I., Rojas, E., and Vergara, J. (1974). Calcium influxes
and tension development in perfused single barnacle muscle
fibers under membrane potential control. J. Physiol.,
243, 523-551.
Axelsson, J., and Thesleff, S.(1959). A study of super-
sensitivity in denervated mammalian skeletal muscle.
J. Physiol., 149,178-193.
Bianchi (1961). The effects of caffeine on radiocalcium movement
in frog sartorius., J. Gen. Physiol., 44, 845-850.
Chiarandini, D.j., Reuben, J.P., Brandt, P.W., and Grundfest, H.
(1970). Effects of caffeine on crayfish muscle fibers.
J. Gen. Physiol., 55, 640-662.
Cohen, J.L., Weiss, K.R., and Kupferman, I.(1978). Motor control
of buccal muscles in Aplysia. J. Neurophysiol., 41,157-180.
Cole, R.A., Twarog, B.M. (1972). Relaxation of catch in a
molluscan smooth muscle. Comp. Bioch. Physiol. 43, 321-330.
Harrison, J. T. (1975). Isometric responses of the somatic
musculature of Cryptochiton stelleri. Veliger 18 (Suppl),
79-82.
Heyer, C.B., Kater, S.B., and Karlsson, U.L.(1973). Neuro-
muscular systems in molluscs. Am. Zool., 13, 247-270.
Hofmann, A. (1969). Caffeine effects on transmitter depletion
and mobilization at motor nerve terminals. Amer. J. Physiol
216, 621-630.
Royle, G. (1983). Nuscles and their neural control. Nev York:
John Wiley and Sons, Inc., 689 pp.
Auddart, M., Hunt,S., and Oates, K. (1977). Calcium movements
during contraction in molluscan smooth muscle, and the
loci of calcium binding and release. J. Exp. Biol. 68,
45-56.
Kobayashi, M., and Muneoka, Y. (1980). Modulatory actions of
octopamine and serotonin on the contraction of buccal
muscles in Rapana thomasiana. I. Enhancement of contraction
in the radula protractor. Comp. Bioch Physiol., 650,73-79.
Mattison, A.G.M., and Arvidsson, J.A.(1966). Some effects of
electrical stimulation and exogenous metabolites on the
contractile activity and the ultrastructure of the radula
muscle of Buccinum undatum, 2. Zellforsch, 73,37-55.
Muneoka, Y., and Mizonishi, T. (1969). Effects of changes in
external calcium concentration on the caffeine contracture
of ABRN of Mytilus edulis. 20ol. Magazine 78, 101-107.
Sandow, A. (1965). Excitation-contraction coupling in skeletal
muscle. Pharm. Rev. 17, 265-320.
Schmidt-Nielsen, J.A., and Weight, F.F. (1976), Synaptic
transmission: long-lasting potentiation by a post-synaptig
mechanism. Science 194,1437-1439.
Iwarog, B.M. (1954). Responses of a molluscan smooth muscle to
acetylcholine and 5-hydroxytryptamine. J. Cell Comp.
Physiol. 44, 141-164.
Twarog, B.M. (1960). Innervation and activity of a molluscan
smooth muscle. J. Physiol. 152,220-235.
Iwarog, B.M. (1967). Excitation of Nytilus smooth muscle.
J. Physiol. 192,857-868.
Twarog, B.M. (1967a). The regulation of catch in molluscan muscle.
J. Gen Physiol. 50, 157-169.
Twarog, B.M. (1968). Possible mechanism of action of serotonin
on molluscan muscle. Adv. Pharmac. 6, 5-15.
Twarog, B.M., and Hidaka, T.(1971). The calcium spike in
Mytilus muscle and the action of serotonin. J. Gen. Physid
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ACKNOWLEDGEMENTS
I thank Hopkins Marine Station for being the great
research facility that it is and for complementing my
Stanford education with unforgettable memories. Very speci
thanks go to my advisors Dr. Mark Denny and Dr. William
Gilly for their guidance and dedicated instruction as well
as to Stuart Thompson for constructive criticism of this
work. To my fiancee Keith Tansey for his patience and
love throughout our Hopkins' days and for a lifetime ahead
FIGURE 1.
FIGURE 2.
FIGURE 3.
FIGURE 4.
FIGURE LEGENDS
Twitch response to single and double stimuli.
The interval between double stimuli is increased
from 2 to 10 seconds.
Summation and Tetanic Response. Increasing the
frequency of repeated stimulation from 1 pulse
per second to 10 pulses per second causes fusion
to a tetanus.
Effects of Solutions and Electrical Stimulation
on the Contraction Stress of the Muscle. The
effect of each solution alone is indicated by
the cross-bars. Additional effects of the first
series of shocks in each solution are indicated
by the clear bars.
Effect of Series of Shocks on Muscle Contraction,
Responses are expressed as net percentage increases
above (or decreases below) the muscle stress evoked
in artificial seawater (ASW), 12 x 10' N/m? and
does not include the loading stress of 9 x 104 N/m2
which was common to all the muscle preparations.
The second series of shocks in the test solutions
are shown'as percentage increases (or decreases)
over the corresponding value in ASW of 11 x 10' N/m2.
The three percentages slightly above the ASW baseline
are in actuality on the baseline denoting a 0% increase.
FIGURE 5. Effect of Caffeine, High K and High Ktcaffeine
on Waveforms and Relaxation Times.
FIGURE 6.
Effect of High K-Caffeine on Summation of
repeated single shocks. High K-Caffeine
potentiates stress accumulation in response
to electrical stimulation. It also interferes
with decrease of muscle stress in the second
application and potentiates subsequent response
to electrical stimuli.
FIGURE
5-Hr Relaxation of Ach-induced contraction stress.
Ach (10""M) bath alone induces a prolonged
contraction (top waveform) that can be relaxed
by 5-HT (10 "M) at a rate 33 times faster than
in Ach.
TABLE 1.
(Adapted from Schmidt-Nielsen 1979). Contraction
Stresses of Various animal muscles. The shell-
foot retractor muscle appears to have a maximum
contraction stress in the lower range of values
listed.
0
6
FIGURE 1
2 Volts 50 ms
Twin Shocks 2 sec delay
2.5 sec delay
5 sec delay
O4
1O sec delay
O1
—
20 25 30
10
15
Time (sec)
at-
O

Qu/NO)s
—-- -
1
8

Q
33
30
27

24.
821
o 18.
15.
12
9.
6
3
FIGURE 3.
ASW 5-HT
L
Caffeine
High K¬
Caffeine
High K
Ach
AoadingTens
Solution alone
Solution shock
)(
)0
1

—
—


2
X
oo
Au

o
S
I
O
OO
(/N)ssa
3
S
4

S



u/NOI) s1
1
+
18
18
1
uN) s
0
TABLE 1.
ANIMAL
Annelid
(Arenicola)
Limpet
(Lottia
gigantea)
Bivalve
(Anodonta)
Bivalve
(Mytilus)
Octopus
Insect
(Locusta)
Insect
(Decticus)
Insect
(Drosophila)
Frog (Rana)
Rabbit
Man
MUSCLE
Body wall
Shell-foot
retractor
Adductor
Anterior byssus
retractor
Eunnel retractor
Hindlegs
Flexor tibiae
Wing muscles
Anterior tibial
"Skeletal muscle'
Ankle flexors
CONTRACTION
STRESS
(N/m2)
3.
5
4.5
5.
4.7
5.9
5
4.4
5.0
4.2
2