Length-dependent activation canine skeletal muscle
of in situ
C. R. LAMBERT, L. B. GLADDEN, AND W. N. STAINSBY Department of Physiology, College of Medicine, University of Florida,
Gainesville,
Florida,
32610
LAMBERT, C. R., L. B. GLADDEN, AND W. N. STAINSBY. Length-dependent activation of in situ canine skeletal muscle. Am. J. Physiol. 237(l): C38-C42, 1979 or Am. J. Physiol.: Cell Physiol. 6(l): C38-C42, 1979.-This study was designed to assess the contribution of length-dependent activation to the peak isometric twitch tension developed and the maximal rate of tension development (dP/dt) of in situ canine skeletal muscle. Length-developed tension and length-dP/dt relationships were generated for the dog gastrocnemius-plantaris muscle group at three different levels of inotropic state as determined by stimulation frequency. These relationships were then normalized with respect to maximal developed tension and maximal dP/dt and the normalized curves were superimposed for comparison. At progressively shorter muscle lengths the augmentation of tension production by a given increment in inotropic state was greater as measured by either developed tension or dP/dt. Thus, a given change in muscle length produced a greater change in performance in less potentiated muscles. These findings are similar to those from studies of isolated cardiac muscle and illustrate the lack of independence between activational state and muscle length for in situ skeletal muscle. skeletal muscle; activation
length-tension
relationship;
length-dependent
YEARS, experimental evidence has accumulated indicating that two separate types of length-dependent modulation interact to influence the magnitude and characteristics of tension production in both skeletal (13) and cardiac muscle (7). The first type of modulation is geometric in nature and depends upon the degree of actin-myosin myofilament overlap in the sarcomere. The second type of modulation depends on a degree of contractile system activation that varies with muscle length and is not simply a function of longitudinal geometric apposition of myofdaments. This latter process of lengthdependent activation has been accepted as a fundamental property of striated muscle and has been incorporated into recent versions of both the crossbridge theory and the electrostatic theory of contraction (9). Most of the experimental work designed to describe the characteristics of length-dependent activation has been done using single fiber preparations of various types in which measurements of sarcomere length are possible. Recently, Lakatta and Jewell (8) have developed a method to assess the contribution of length-dependent activation to the isometric contraction of intact feline papillary muscle. Their approach involves obtaining length-tension curves for muscles at different levels of
IN RECENT
C*38
activation as determined by inotropic state. The inotropic state was varied by altering the rate and pattern of stimulation. The curves obtained are then normalized with respect to maximal developed tension and compared. If the inotropic state or activational state and muscle length are independent regulators of tension generation, the normalized curves should be superimposable. If the inotropic or activational state and muscle length are not independent in their effects on tension production, the normalized curves should differ. The present study was designed to ascertain whether length and inotropic state are independent regulators of tension generation in isometric twitch contractions of in situ canine skeletal muscle. The experiments were designed around the method developed by Lakatta and Jewell (8). METHODS
Mongrel dogs of either sex were anesthetized with sodium pentobarbital (30 mg/kg iv); additional doses were given as needed during the experiments. The animals were intubated and allowed to breath spontaneously. The left gastrocnemius-plantaris muscle group was isolated and the Achilles tendon was sectioned and secured in an angle aluminum clamp for attachment to the muscle lever that has been described previously (14). The sciatic nerve was isolated close to the muscle and was stimulated with supramaximal rectangular voltage pulses, 0.2 ms in duration, supplied by a Grass SD9 stimulator, isolated from ground. A record of muscle tension, the first derivative of tension with respect to time (dP/dt), and muscle length were obtained by an oscillographic recorder with a frequency response flat to 50 Hz (Astro-Med). The differentiator was calibrated at the conclusion of each experiment with ramp voltages from a function generator (Wavetek). The force transducer was calibrated by direct suspension of weights from the muscle lever and the length recording was calibrated by moving the lever against a scale at the point of muscle attachment. Following a brief period of equilibration, optimal muscle length (L,) was determined by progressively lengthening the muscle using the lever preload set screw and monitoring developed tension. The actual length of the muscle at L, was measured to the nearest mm at the end of an experiment. After determination of L, the muscle was shortened to a length where developed tension was approximately 4040% of that at L,. At this muscle length
0363~6143/79/0000-OOOOooo$Ol.25
Copyright
0 1979 the American
Physiological
Society
Downloaded from www.physiology.org/journal/ajpcell by ${individualUser.givenNames} ${individualUser.surname} (129.081.226.078) on January 15, 2019.
LENGTH-DEPENDENT
c39
ACTIVATION
a control period was allowed with a stimulation frequency of 0.2 Hz until steady-state levels of tension and dP/dt were achieved. The stimulation frequency was then increased to 1.0Hz and 2.0 Hz with a period of time allowed to reach a steady state at each frequency. The muscle was then lengthened an increment judged to be approximately 33% of the distance between L, and the length where developed tension was 40,50% of that at L,. Following the length step, the frequency sequence was repeated allowing time to obtain steady-state tension and dP/dt measurements. This procedure was repeated following two more length steps so that tension and dP/dt data were obtained at four muscle lengths including Lo. Developed tension was normalized with respect to muscle wet weight determined after excision at the end of the experiment. This method of normalization has been widely used in order to avoid problems inherent in attempting an extrapolation to cross-sectional area due to the penate geometry of the gastrocnemius (14). The muscle lengths at which measurements were taken were normalized with respect to the measured value of L,. Statistical comparisons were made using Student’s t test for paired data. The results presented below were obtained from experiments on four muscles.
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of in situ
C. R. LAMBERT, L. B. GLADDEN, AND W. N. STAINSBY Department of Physiology, College of Medicine, University of Florida,
Gainesville,
Florida,
32610
LAMBERT, C. R., L. B. GLADDEN, AND W. N. STAINSBY. Length-dependent activation of in situ canine skeletal muscle. Am. J. Physiol. 237(l): C38-C42, 1979 or Am. J. Physiol.: Cell Physiol. 6(l): C38-C42, 1979.-This study was designed to assess the contribution of length-dependent activation to the peak isometric twitch tension developed and the maximal rate of tension development (dP/dt) of in situ canine skeletal muscle. Length-developed tension and length-dP/dt relationships were generated for the dog gastrocnemius-plantaris muscle group at three different levels of inotropic state as determined by stimulation frequency. These relationships were then normalized with respect to maximal developed tension and maximal dP/dt and the normalized curves were superimposed for comparison. At progressively shorter muscle lengths the augmentation of tension production by a given increment in inotropic state was greater as measured by either developed tension or dP/dt. Thus, a given change in muscle length produced a greater change in performance in less potentiated muscles. These findings are similar to those from studies of isolated cardiac muscle and illustrate the lack of independence between activational state and muscle length for in situ skeletal muscle. skeletal muscle; activation
length-tension
relationship;
length-dependent
YEARS, experimental evidence has accumulated indicating that two separate types of length-dependent modulation interact to influence the magnitude and characteristics of tension production in both skeletal (13) and cardiac muscle (7). The first type of modulation is geometric in nature and depends upon the degree of actin-myosin myofilament overlap in the sarcomere. The second type of modulation depends on a degree of contractile system activation that varies with muscle length and is not simply a function of longitudinal geometric apposition of myofdaments. This latter process of lengthdependent activation has been accepted as a fundamental property of striated muscle and has been incorporated into recent versions of both the crossbridge theory and the electrostatic theory of contraction (9). Most of the experimental work designed to describe the characteristics of length-dependent activation has been done using single fiber preparations of various types in which measurements of sarcomere length are possible. Recently, Lakatta and Jewell (8) have developed a method to assess the contribution of length-dependent activation to the isometric contraction of intact feline papillary muscle. Their approach involves obtaining length-tension curves for muscles at different levels of
IN RECENT
C*38
activation as determined by inotropic state. The inotropic state was varied by altering the rate and pattern of stimulation. The curves obtained are then normalized with respect to maximal developed tension and compared. If the inotropic state or activational state and muscle length are independent regulators of tension generation, the normalized curves should be superimposable. If the inotropic or activational state and muscle length are not independent in their effects on tension production, the normalized curves should differ. The present study was designed to ascertain whether length and inotropic state are independent regulators of tension generation in isometric twitch contractions of in situ canine skeletal muscle. The experiments were designed around the method developed by Lakatta and Jewell (8). METHODS
Mongrel dogs of either sex were anesthetized with sodium pentobarbital (30 mg/kg iv); additional doses were given as needed during the experiments. The animals were intubated and allowed to breath spontaneously. The left gastrocnemius-plantaris muscle group was isolated and the Achilles tendon was sectioned and secured in an angle aluminum clamp for attachment to the muscle lever that has been described previously (14). The sciatic nerve was isolated close to the muscle and was stimulated with supramaximal rectangular voltage pulses, 0.2 ms in duration, supplied by a Grass SD9 stimulator, isolated from ground. A record of muscle tension, the first derivative of tension with respect to time (dP/dt), and muscle length were obtained by an oscillographic recorder with a frequency response flat to 50 Hz (Astro-Med). The differentiator was calibrated at the conclusion of each experiment with ramp voltages from a function generator (Wavetek). The force transducer was calibrated by direct suspension of weights from the muscle lever and the length recording was calibrated by moving the lever against a scale at the point of muscle attachment. Following a brief period of equilibration, optimal muscle length (L,) was determined by progressively lengthening the muscle using the lever preload set screw and monitoring developed tension. The actual length of the muscle at L, was measured to the nearest mm at the end of an experiment. After determination of L, the muscle was shortened to a length where developed tension was approximately 4040% of that at L,. At this muscle length
0363~6143/79/0000-OOOOooo$Ol.25
Copyright
0 1979 the American
Physiological
Society
Downloaded from www.physiology.org/journal/ajpcell by ${individualUser.givenNames} ${individualUser.surname} (129.081.226.078) on January 15, 2019.
LENGTH-DEPENDENT
c39
ACTIVATION
a control period was allowed with a stimulation frequency of 0.2 Hz until steady-state levels of tension and dP/dt were achieved. The stimulation frequency was then increased to 1.0Hz and 2.0 Hz with a period of time allowed to reach a steady state at each frequency. The muscle was then lengthened an increment judged to be approximately 33% of the distance between L, and the length where developed tension was 40,50% of that at L,. Following the length step, the frequency sequence was repeated allowing time to obtain steady-state tension and dP/dt measurements. This procedure was repeated following two more length steps so that tension and dP/dt data were obtained at four muscle lengths including Lo. Developed tension was normalized with respect to muscle wet weight determined after excision at the end of the experiment. This method of normalization has been widely used in order to avoid problems inherent in attempting an extrapolation to cross-sectional area due to the penate geometry of the gastrocnemius (14). The muscle lengths at which measurements were taken were normalized with respect to the measured value of L,. Statistical comparisons were made using Student’s t test for paired data. The results presented below were obtained from experiments on four muscles.
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