Temperature sensitivity of tension development in a

Isometric contractions of the biceps brachii (short head) muscle in the rat were recorded in vitro with direct stimulation and at different temperatures (range, 35°C-10°C). In confirmation of our previous findings from fast extensor digitorum longus and slow soleus muscles, the time and rate parameters of the twitch and the tetanus showed an increased temperature sensitivity below 20°C.The dependence on the initial muscle length of the rate of rise of tetanic tension was examined at 27°C and at 15°C.When represented as a percentage of the tetanic tension at each length, the rate of rise was independent of muscle length at both temperatures. Our interpretation of this particular observation is that the increased cooling depression of the rate of tension rise below 20°C is not associated with a qualitative change in its underlying basis. MUSCLE 81 NERVE

7:298-303

1984

TEMPERATURE SENSITIVITY OF TENSION DEVELOPMENT IN A FAST-TWITCH MUSCLE OF THE RAT MURTADA H. ELMUBARAK, MBBS, and K. W. RANATUNGA. PhD

Several contraction parameters of mammalian on the initial muscle length of the rate of tension skeletal muscle exhibit an increased temperature rise was examined at 27°C and at 15°C. sensitivity when cooled below 20"C-25"C.s~"10~'2~1~ Consequently, the temperature coefficients (Qlo) METHODS determined for temperatures below 20°C were The experiments reported in this article were carconsiderably higher than those obtained for temried out on the short head (smaller of the two peratures above 25°C. This was shown to be true heads) of the biceps brachii muscle, which was for the maximum rate of rise of isometric tetanic isolated from male rats of' about 4 weeks of age. tension in the rat f'ast- and slow-twitch A rat was anesthetized with an intraperitoneal Such a situation could arise if qualitatively differinjection (50 mg/kg of body weight) of sodium ent processes underlie the tension rise at the two pentobarbitone (Sagatal, May & Baker Ltd., Dagtemperature range^.^ It was, therefore, of interest enham, England) and the whole biceps brachii to examine the relation of the rate of tension rise to muscle complex was dissected out. Separation of some other independent variable at a temperature the short head of the biceps brachii was done in a higher than 25°C and at a temperature lower than perspex chamber filled with physiological saline so20°C. We have carried out a similar experiment on lution. T h e preparation was then set up horizonthe biceps brachii muscle, where the dependence tally in a muscle chamber for in vitro isometric tension recording, as described in previous articles."," T h e saline solution contained 115 mM NaC1, 5 From the Department of Physiology, The Medical School, University of Bristol, Bristol, England. mM KCl, 4 mM CaC12, 1 mM MgC12, 1 mM Na H2P04, 11 mM glucose, 24 mM NaHC03, and 10Acknowledgments: M. H. Elmubarak is in receipt of a postgraduate scholarship from the University of Khartoum in Sudan. We are grateful to 20 mg/l of tubocurarine chloride. The saline Mrs. A. Lear for typing the manuscript. was continuously bubbled with a mixture of 95% Address reprint requests to Dr. Ranatunga at the Department of Physiol0 2 and 5% CO2 and was replaced at a rate of about ogy, The Medical School, University of Bristol, Bristol 858 ITD, England. 1-2 ml/minute. The pH of the saline in the musReceived for publication June 17, 1983; revised manuscript accepted for cle chamber was monitored and found to be within publication October 20,1983. 7.2 and 7.6 in different experiments and at differ0148-639X/0704/0298 $04.0010 0 1984 John Wiley & Sons, Inc. ent temperatures.

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May 1984

Detailed information about stimulation, recording tension and rate of tension rise, control, and variation of temperature of the saline in the muscle chamber has been previously reported.9 Since rat biceps brachii muscle has not been previously used in this type of experiment, a series of experiments were carried out to confirm the findings previously made on extensor digitorum longus and soleus muscles.'"v' The effect of temperature on the isometric twitch contraction (peak tension, time-to-peak, and time-tohalf relaxation) and on the isometric tetanic contraction (plateau tension and rate of tension rise) was examined by a procedure essentially similar to that adopted in the previous study." Isometric contractions were examined at 35°C and at 5°C intervals in cooling from 35°C to 10°C and during rewarming from 10°C to 35°C. T h e frequencies and durations of tetanic stimuli required for eliciting fused tetani were determined in preliminary experiments and were found to be very similar to those used for the extensor digitorum longus muscle.""' In a second series of experiments on five muscles, the dependence on the muscle length of the tetanic tension and the rate of tension rise was examined at 27°C and at 15°C. T h e muscle length was first adjusted to that at which twitch tension was maximal and a tetanic contraction was recorded. Tetanic contractions were then recorded at progressively longer muscle lengths (set by micrometer on which the tension transducer was mounted) until the tension dropped to, in some cases, 25% of the maximal. A series of recordings were then made when the length was progressively decreased until the tension at a short length was around 25% of the maximal. A final series of recordings were made as the length was increased back to o r beyond the optimal length range. The frequency of tetanic stimulation was 300-350 Hz at 27°C and 100-150 Hz at 15"C, and they resulted in smooth and fused differentiated tension records. The durations were 75 msec and 150 msec, respectively, at 27°C and 15°C. The resting (passive tension) was electronically subtracted from the tension transducer output' and the resting tension, the tetanic tension, and the positive peak of the differentiated tension record (equal to the maximum rate of tension rise) were measured from each c o n t r a ~ t i o n .T~h e cycle of stimulation adopted was once every 1-1.5 minutes and tetanic

Plan of the Study.

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Temperature on Muscle Contraction

contractions were not elicited at intervals shorter than 4 minutes. At the end of a recording session, a muscle length was measured in the chamber and it was fixed at a known length setting in relation to the length-tension relationship. Fixation was done by replacing saline in the chamber with 50% ethyl alcohol for about 5- 10 minutes. The stiffened muscle was then carefully transferred to another chamber, pinned at the same length, and fixed overnight in formol-saline solution. The muscles were macerated in 25% nitric acid and stored in 50% glycerol. Between 6 and 10 muscle fibers, or bundles of muscle fibers containing up to about 6 fibers, were teased out from each muscle and their sarcomere lengths were sequentially determined along 80%-90% of their length by using a laser diffraction technique. Muscle fiber lengths were determined by microscopical examination ( x 100 magnification) in the same preparations and in additional fiber bundles (1 1-28 bundles per muscle) containing 10-20 fibers in each. Lengths of fibers within a bundle did not vary by more than 0.2 mm. Using these measurements, the mean muscle fiber length and the mean sarcomere length were determined for each muscle. These determinations were employed in the construction of the sarcomere length axis for each muscle. The procedure used in histological processing is essentially similar to that described by Close.' The rate and time (as reciprocals) parameters recorded at different temperatures were examined in the form of Arrhenius plots; i.e., by plotting the logarithm of a parameter against the reciprocal absolute temperature. Since rigorous Arrhenius analyses may not be valid for these measurements, the temperature sensitivity will be expressed as Qlo. These were calculated from the regressions fitted to the pooled data, for 35°C25°C and for 20°C-10°C.

Analysis.

RESULTS AND DISCUSSION

The short head of the biceps brachii muscle in the rat is a small muscle compared to the soleus and the extensor digitorum longus (EDL) muscles used in our previous experiments. It has discrete tendons of origin and insertion-the latter being shared with the long head. Its muscle fibers are nearly parallel to one another. In a given muscle, more than 95% of the muscle fibers had lengths which were within & 10% of the average muscle fiber length. At near optimal length, the ratio of

MUSCLE & NERVE

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299

~

~

~

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_

_

_

Table 1. Summary data for the isometric contraction measurements from the short head of the biceps brachii muscle in 4-week-old male rats * ~

Time-to-peak twitch tension (msec)

Time-to-half relaxation (msec)

9.5 t 0.5 (10)

10.7

?

(10)

0.4

~

~

~

~

~

Twitch-tetanus tension ratio

Rate of tension rise

Cooling potentiation

0 19 t- 0.01 (10)

4.5 2 0.04 ( 8)

7.65 t- 0.08 (5)

~

*Recordings were made at 35°C in vitro and each value gives the mean & SEM with the number of muscles in parentheses. The rate of tetanic tension rise is given as a percentage of tetanic tension per millisecond ("7 P,/msec) and the cooling potentiation is the ratio of the twitch tension at 20°C to the average twitch tension recorded at 35°Cprior to cooling and after rewarming.

average muscle fiber length to muscle length was 0.6 0.02 (mean & SEM, n = 5). The ratios for soleus and EDL muscles were 0.64 -+ 0.02 ( n = 6) and 0.43 & 0.01 (Q = 7), respectively (Ranatunga and Wylie, 1982, unpublished observations). As the mean data in Table 1 show, the time course of the isometric twitch, the ratio of twitch tension to tetanic tension, and the maximum rate of tetanic tension rise were comparable to those of the EDL muscle. Additionally, cooling from 35°C to 20°C potentiated the twitch tension (see the last column in Table 1) as in the fast-twitch EDL muscle.8 These findings establish that this muscle contains predominantly fast-twitch muscle fibers. Insofar as it had less tendon compliance (see the preceding), the short head of the biceps brachii muscle is perhaps better suited for this type of experiment than the fast EDL muscle previously used. T h e temperature dependence for several contraction parameters (time-to-peak twitch, time-tohalf relaxation of twitch, and the rate of rise of tetanic tension), when examined in the form of Arrhenius plots,' showed clear evidence of an increase of temperature sensitivity in cooling below about 23°C. Figure 1 is an Arrhenius plot of the maximum rate of tetanic tension rise, represented as a percentage of the tetanic tension recorded at each temperature (i.e., % Po/msec). The two straight lines represent the calculated regressions ( r > 0.6, n > 19) for temperatures higher than 23°C and lower than 23°C. The Qlo determined from the regressions were 1.48 and 2.75, respectively, for higher (35OC-25"C) and lower (20°C10°C) temperature ranges. Similar analyses on the time-to-peak and the time-to-half relaxation in the twitch also showed higher Q l o for the lower temperature range. These results confirm our previous findings made on different muscle preparations.' The length dependence of tension and rate of rise of tension in isometric tetani was examined in experiments on five muscles in which nine rela-

*

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300

Temperature on Muscle Contraction

tions were analyzed. Figure 2 shows experimental records from two biceps brachii muscle preparations (upper row and lower row) at 27°C. Each frame contains an isometric tetanus and the differentiated tension record, the positive peak of which was measured as the maximum rate of tension rise. T h e records from each muscle were obtained at different initial lengths. The numbers given above each frame denote, in millimeters, the difference in length setting (in relation to the length setting at the higher end of optimal length range). Thus, in the upper row, the middle record was taken at the optimal length range and the left and the right records, respectively, were taken at shorter and longer muscle lengths. In the lower row, the third record from the left was at an optimal length range (but taken at half the gain to avoid amplifier overloading); the two records on the left side were at shorter lengths and the record on the right side was at a longer length. The records show that, to a first approximation, there is a

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(11~1.10~ FIGURE 1. An Arrhenius plot of the maximum rate of tension rise in isometric tetanus. The rate is plotted along a logarithmic ordinate and reciprocal of the absolute temperature along the abscissa (labeled also in "C). Data are from four muscles in which each symbol represents the mean and the vertical lines represent the 2 SEM. Results include data obtained in cooling (half-filled circle) and rewarming (open circle).

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May 1984

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