Comparison between the effect of static contraction and tendon stretch on the discharge of group III and IV muscle afferents

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1 J Appl Physiol 99: , First published June 30, 2005; doi: /japplphysiol Comparison between the effect of static contraction and tendon stretch on the discharge of group III and IV muscle afferents Shawn G. Hayes, Angela E. Kindig, and Marc P. Kaufman Division of Cardiovascular Medicine, University of California, Davis, California Submitted 26 May 2005; accepted in final form 29 June 2005 Hayes, Shawn G., Angela E. Kindig, and Marc P. Kaufman. Comparison between the effect of static contraction and tendon stretch on the discharge of group III and IV muscle afferents. J Appl Physiol 99: , First published June 30, 2005; doi: /japplphysiol The exercise pressor reflex is evoked by both mechanical and metabolic stimuli. Tendon stretch does not increase muscle metabolism and therefore is used to investigate the mechanical component of the exercise pressor reflex. An important assumption underlying the use of tendon stretch to study the mechanical component of the exercise pressor reflex is that stretch stimulates the same group III mechanosensitive muscle afferents as does static contraction. We have tested the veracity of this assumption in decerebrated cats by comparing the responses of group III and IV muscle afferents to tendon stretch with those to static contraction. The tension-time indexes as well as the peak tension development for both maneuvers did not significantly differ. We found that static contraction of the triceps surae muscles stimulated 18 of 30 group III afferents and 8 of 11 group IV afferents. Similarly, tendon stretch stimulated 14 of 30 group III afferents and 3 of 11 group IV afferents. However, of the 18 group III afferents that responded to static contraction and the 14 group III afferents that responded to tendon stretch, only 7 responded to both stimuli. On average, the conduction velocities of the 18 group III afferents that responded to static contraction ( m/s) were significantly slower (P 0.03) than those of the 14 group III afferents that responded to tendon stretch ( m/s). We have concluded that tendon stretch stimulated a different population of group III mechanosensitive muscle afferents than did static contraction. Although there is some overlap between the two populations of group III mechanosensitive afferents, it is not large, comprising less than half of the group III afferents responding to static contraction. exercise pressor reflex; thin fiber mechanoreceptors; cats; autonomic nervous system; control of the circulation THE EXERCISE PRESSOR REFLEX is evoked by contraction of skeletal muscle and consists of increases in mean arterial blood pressure, heart rate, and breathing (17). Group III and IV muscle afferents are believed to comprise the afferent arm of the exercise pressor reflex arc (12). Contraction of skeletal muscle generates two types of stimuli that activate these thinly myelinated (i.e., group III) and unmyelinated (i.e., group IV) afferents. The first stimulus is mechanical and occurs when contraction distorts the receptive fields of the afferents. The second is metabolic and occurs when contraction generates chemical by-products. For the most part, group III afferents respond to mechanical stimuli and group IV afferents respond to metabolic stimuli (7 9). Nevertheless, the sensitivity of group III afferents to mechanical stimuli is increased when the concentrations of metabolic by-products of contraction are increased (20, 23). Similarly, the sensitivity of group III afferents to mechanical stimuli is decreased when these concentrations are decreased (20, 23). Consequently, contraction of skeletal muscle is not a useful stimulus to isolate the mechanical component of the exercise pressor reflex. Instead, tendon stretch is often used to study the mechanical component of the reflex because it does not generate any metabolites in the muscle (4, 25, 28). An important assumption underlying the use of tendon stretch to study the mechanical component of the exercise pressor reflex is that tendon stretch stimulates the same group III mechanoreceptors as does contraction. Another assumption is that the magnitude of the responses of these mechanoreceptors is the same for both stretch and contraction. In the experiments to be described, we have examined the veracity of these assumptions by comparing the responses of group III and IV muscle afferents to contraction with those to tendon stretch. METHODS General. The Institutional Care and Use Committee of the University of California, Davis approved all procedures employed in this report. Anesthesia was induced in cats (n 38; kg) with 5% halothane in oxygen. The trachea was cannulated, and the lungs were ventilated mechanically (Harvard Apparatus) with 3% halothane in oxygen until the end of surgery. Catheters were placed in the right jugular vein and common carotid artery for delivery of drugs and for measurement of arterial blood pressure, respectively. The carotid artery catheter was connected to a pressure transducer (model P23 XL, Statham). Heart rate was calculated on a beat-to-beat basis from the arterial pressure pulse (Gould Biotech). The cat was placed in a Kopf stereotaxic and spinal unit and then given dexamethasone (4 mg iv). A midcollicular decerebration was performed, and all neural tissue rostral to the section was removed. Bleeding was controlled, and the cranial vault was filled with agar (37 C). A laminectomy was performed that exposed the fourth lumbar through second sacral segments. The triceps surae muscles of the left leg were isolated, and the calcaneal bone was cut. The knee was bent at an angle of 115. The free end of the calcaneal tendon was attached to a force transducer (model FT-10C, Grass) to measure tension development when the triceps surae muscles were either contracted statically or were stretched. All visible branches of the left sciatic nerve innervating the thigh and hip as well as the left femoral nerve were cut. The anesthesia was terminated after all the surgery was completed. Recording of impulse activity from group III and IV afferents. The impulse activity from group III and IV afferents with endings in the left triceps surae muscles was recorded from fine filaments dissected from the L 7 or S 1 dorsal roots. The peripheral cut ends of these filaments were placed on one foot of a bipolar hook electrode. The other foot was grounded to the cat with a thin string soaked in saline. Address for reprint requests and other correspondence: S. G. Hayes, Div. of Cardiovascular Medicine, One Shields Dr., Univ. of California, Davis, CA ( sghayes@ucdavis.edu). The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact /05 $8.00 Copyright 2005 the American Physiological Society 1891

2 1892 GROUP III RESPONSES TO CONTRACTION AND STRETCH The afferent signals were passed through a high-impedance probe (model HIP 511, Grass Instruments), amplified and filtered (100 3,000 Hz; model P 511, Grass Instruments). Action potentials were displayed on a computer monitor (Spike 2, Cambridge Electronics Design, Cambridge, UK) and on a storage oscilloscope (model HP 54603B). The conduction velocity of an afferent was calculated by dividing the conduction distance between the recording electrode on the dorsal root and the stimulating electrode on the tibial nerve by the conduction time, which was measured on the storage oscilloscope. Afferents conducting impulses between 2.5 and 30 m/s were classified as group III afferents, and those conducting impulses at 2.5 m/s were classified as group IV afferents (7). The receptive fields of the afferents were located in the triceps surae muscles by either gently stroking the muscle, by squeezing the muscle in a nonnoxious manner, or by pinching the muscle in a noxious manner. Afferents conducting impulses faster than 30 m/s were discarded. The triceps surae muscles were contracted statically for 60 s by electrical stimulation (15 25 Hz) of either the cut peripheral ends of the L 7 and S 1 ventral roots or the intact tibial nerve. In the former case, the pulse width was 0.1 ms, and in the latter case the pulse width was 25 s. The current applied to both the ventral roots and the tibial nerve was times the threshold current needed to twitch the triceps surae muscles. The triceps surae muscles were stretched for 60 s by turning a rack and pinion that was attached to the calcaneal tendon. While performing the experiments, we attempted to match the magnitudes of the tension traces for static contraction and tendon stretch. The order of presentation of the two stimuli was varied randomly. Data analysis. The baseline impulse activity of each group III and IV afferent was counted for the 60-s period immediately preceding either static contraction or tendon stretch. Similarly, the impulse activity of the afferent was counted for the 60-s period comprising either static contraction or tendon stretch. Both counts were divided by 60 so that they could be expressed as impulses per second. Our criterion for stimulation of a group III or IV afferent by either tendon stretch or static contraction was selected a priori and consisted of an increase in activity of 12 impulses in 60 s. The TTI (19) for both static contraction and tendon stretch was calculated by integrating the area between the tension trace and its baseline level (Spike 2). Peak developed tension was calculated by subtracting baseline tension from peak tension. All values are expressed as means SE. Paired t-tests or two-way repeated-measures ANOVA followed by Tukey s post hoc tests were used to determine statistical significance. The criterion for statistical significance was set at P RESULTS We recorded the impulse activity of 30 group III afferents with receptive fields in the triceps surae muscles. Five of these 30 group III afferents did not respond to either static contraction or tendon stretch and were consequently discarded. Each of the remaining 25 group III afferents (conduction velocity m/s; range: m/s) responded to either tendon stretch or static contraction or both, and they formed the basis of our analysis. Static contraction of the triceps surae muscles for 60 s stimulated 18 of the 25 group III afferents. Similarly, tendon stretch for 60 s stimulated 14 of the 25 group III afferents (Figs. 1 and 2). Although tendon stretch stimulated fewer group III afferents than did static contraction, five of the group III afferents classified as nonresponsive to stretch did show small increases in activity above baseline levels. These increases were in the range of 5 10 impulses over the 60 s of tendon stretch (Fig. 3). A comparison between the responses of the 25 group III afferents to contraction and the responses to tendon stretch Fig. 1. Effects of tendon stretch (TS) and static contraction (CX) on the discharge of group III and IV muscle afferents. Values are means SE; n, no. of cats. Open bars, values during baseline (Base); filled bars, values during either tendon stretch or static contraction. Imp, impulses. *Significant difference between baseline and its corresponding value during either tendon stretch or static contraction, P Horizontal bracket represents a significant difference between the magnitude of the responses of the group IV afferents to tendon stretch and the magnitude of the responses of the group IV afferents to static contraction, P revealed that contraction had a similar effect on the average discharge of these afferents as did tendon stretch (Fig. 1). The TTIs did not differ significantly (P 0.38) and averaged kg s for stretch and kg s for contraction. Similarly, peak developed tension did not differ (P 0.79) and averaged kg for stretch and kg for contraction. However, of the 18 group III afferents that responded to static contraction (conduction velocity m/s), only 7 responded to tendon stretch (conduction velocity m/s; Fig. 3). The conduction velocities of the remaining 11 group III afferents that responded to static contraction but not to tendon stretch averaged m/s. The TTIs for the two maneuvers did not differ significantly (P 0.39), averaging kg s for stretch and kg s for contraction. Similarly, peak developed tensions did not differ (P 0.96), averaging kg for stretch and kg for contraction. Of the 14 group III afferents that responded to tendon stretch (conduction velocity m/s), only 7 responded to

3 GROUP III RESPONSES TO CONTRACTION AND STRETCH 1893 stretch ( m/s) were significantly faster (P 0.03) than the conduction velocities of the 18 group III afferents responding to static contraction ( m/s). We also recorded the impulse activity of 11 group IV afferents with endings in the triceps surae muscles (conduction velocity m/s; range: m/s). Static contraction of the triceps surae muscles for 60 s stimulated 8 of the 11 group IV afferents, whereas tendon stretch for 60 s stimulated only 3 of the 11 group IV afferents; these 3 also responded to contraction. Static contraction had a significantly greater (P 0.02) effect on the discharge of the 11 group IV afferents than did tendon stretch (Fig. 1), the latter having no significant effect (P 0.11; n 11). DISCUSSION Fig. 2. Time course of the responses of the group III afferents responding to static contraction (top) and to tendon stretch (bottom). Each maneuver lasted for 60 s and started at time 0. Values are means SE; n, no. of cats. F, Discharge rate calculated for 2 s. CV, conduction velocity. static contraction (conduction velocity m/s; Figs. 4 and 5). The TTIs for the two maneuvers did not differ significantly (P 0.34) and averaged kg s for stretch and kg s for contraction. Similarly, peak developed tension for the two maneuvers did not differ significantly (P 0.26) and averaged kg for stretch and kg for contraction. In addition, the conduction velocities of the 14 group III afferents responding to tendon When considered as a population, group III muscle afferents responded equally to both tendon stretch and static contraction in our experiments. When considered individually, however, group III afferents frequently responded to one stimulus but not the other. Specifically, tendon stretch stimulated only 7 of the 18 group III afferents responsive to static contraction. Similarly, contraction stimulated only 7 of the 14 group III afferents responsive to tendon stretch. In addition, group IV muscle afferents, on average, responded to static contraction but not to tendon stretch. The exercise pressor reflex is widely recognized to be evoked by two general types of stimuli, namely mechanical and metabolic (6, 8). Historically, afferent input from exercising skeletal muscle was believed to signal the central nervous system that blood and/or oxygen supply to the metabolically active tissues was not adequate to meet blood and/or oxygen demand (16, 21, 22). Consequently, metabolic stimuli providing an error signal that blood supply and demand in contracting muscle were not matched properly became the primary focus of investigations into the exercise pressor reflex. Recently, however, mechanical stimuli evoking the exercise pressor reflex have received some attention, especially because they might play an important role in regulating the cardiovascular Fig. 3. Static contraction but not tendon stretch stimulated a group III afferent (conduction velocity: 4.7 m/s). Both maneuvers started at time 0 and lasted for 60 s. Top: plots of the discharge rate of the group III afferent calculated every 2 s. Insets in top: actual recording of impulse activity at the time point shown in the corresponding arrow pointing downward. Horizontal bars, 250 ms; upward-pointing arrowhead, start of tendon stretch. In inset for static contraction, asterisks have been placed over action potentials. Large evenly spaced vertical lines are stimulus artifact. Bottom: plots of the tension developed calculated in 2-s intervals. TTI, tension-time index.

4 1894 GROUP III RESPONSES TO CONTRACTION AND STRETCH Fig. 4. Tendon stretch, but not static contraction, stimulated a group III afferent (conduction velocity: 15.9 m/s). Both maneuvers started at time 0 and lasted for 60 s. Top: plots of the discharge rate of the group III afferent calculated every 2 s. Insets in top: recording of impulse activity at the time point shown in the corresponding arrow pointing downward. Horizontal bar in insets, 250 ms; upward-pointing arrowhead, start of tendon stretch. In inset for static contraction, asterisks have been placed over each action potential. Large evenly spaced vertical lines are the stimulus artifact. Bottom: plots of the tension developed calculated in 2-s intervals. system during exercise in patients (14, 15) and animals with heart failure (11). Mechanical stimuli such as tendon stretch and light, nonnoxious probing of receptive fields have been known for some time to stimulate group III muscle afferents (7, 13, 18). In addition, tendon stretch has been shown to reflexly increase arterial pressure and heart rate in animals (4, 25, 28) as well as heart rate in humans (3). Moreover, compression of muscles in humans, a mechanical stimulus, has also been shown to increase reflexly arterial pressure and heart rate (2, 27). Tendon stretch in animals was reported to not increase metabolism in the muscle (25), and in humans it was reported not to be painful (3). The levels of static contraction and tendon stretch used in our experiments were moderate, most likely averaging less than one-half of maximum. We used relatively low levels of contraction and stretch to minimize nociceptive stimulation of the group III and IV afferents. Despite this precaution, we can offer no assurance that the stimuli used in our experiments were not noxious. For example, Stebbins et al. (25) measured the developed tension evoked when stretching the triceps surae muscles of cats within and beyond their physiological range. They found that a developed tension of 2.4 kg exceeded the physiological range for muscle length. The tensions developed during tendon stretch in our experiments exceeded this level, clearly raising the possibility that they were noxious at least to some degree. Nevertheless, the tension levels generated by either contraction or stretch in our experiments were considerably lower than those used by others. For example, the peak tensions developed by either static contraction or tendon stretch reported by Leshnower et al. (10) averaged approximately twice those reported by us in the present study. In our experiments, the conduction velocities of the group III afferents stimulated by tendon stretch were significantly higher Fig. 5. Static contraction and tendon stretch stimulated a group III afferent (conduction velocity: 17.2 m/s). Top: plots of the discharge rate of the group III afferent calculated every 2 s. Insets in top: actual recording of impulse activity at the time point shown in the corresponding arrow pointing downward. Horizontal bar in insets, 250 ms. In inset for static contraction, asterisks have been placed over each action potential. Large evenly spaced vertical lines are from the stimulus artifact. Bottom: plots of the tension developed calculated in 2-s intervals.

5 on average than those of the group III afferents stimulated by static contraction. This finding may have important functional implications. Specifically, in anesthetized cats, Coote and Pérez-González (1) found that electrical stimulation of group III hindlimb muscle afferents with axons conducting impulses at 15 m/s reflexly decreased sympathetic discharge, whereas stimulation of group III afferents with axons conducting impulses at 15 m/s reflexly increased sympathetic discharge. Consequently, the possibility exists that some of the more rapidly conducting group III afferents stimulated by tendon stretch in our experiments had an inhibitory effect on the discharge of some sympathetic neurons. We can offer no explanation for our observation that some group III afferents responded solely to lengthening the muscle (i.e., tendon stretch), whereas others responded solely to shortening the muscle (i.e., static contraction). Clearly both were mechanosensitive, but their endings, like muscle spindles (i.e., group Ia and II afferents), appear to be situated in parallel with the muscle fibers in which they are embedded. Unlike muscle spindles, group III afferents have free nerve endings that are found frequently near or in collagen (26). Furthermore, group III afferents are much less sensitive to stretch than are spindles. We used two methods to quantify the amount of tension developed by the triceps surae muscles when they were either contracted statically or were stretched. The first, peak developed tension, is commonly used but does not take into account factors such as fatigue or stretch relaxation that causes the muscles to decrease their tension development. The second, TTI, integrates the area under the tension curve and has been shown to correlate strongly with the magnitude of the reflex pressor response to static contraction (19). Both methods of quantification yielded the same result, namely that in our experiments the stimulus generated by static contraction did not differ significantly from the stimulus generated by tendon stretch. In summary, the purpose of our experiments was to test two assumptions. The first was that tendon stretch stimulated the same group III mechanoreceptors as did static contraction. The second assumption was that the magnitudes of the responses of group III mechanoreceptors to static contraction and to tendon stretch were the same. Evidence favoring both assumptions would validate tendon stretch as a useful method of studying the role played by group III mechanoreceptors in evoking the exercise pressor reflex. The mechanical component of this reflex has been shown to be present in both humans and animals and can be quite substantial (3 5, 14, 15, 24). With respect to the first assumption, our findings have shown that tendon stretch stimulated a different population of group III mechanoreceptors than did static contraction. Although there is some overlap in that stretch and contraction stimulate some of the same group III mechanoreceptors, this overlap is not large, comprising less than one-half of the group III mechanoreceptors responsive to static contraction. With respect to the second assumption, the magnitude of the responses of the group III mechanoreceptors responding to tendon stretch was similar to the magnitude of the responses of the group III mechanoreceptors responding to static contraction (see Figs. 1 and 2), even if they were often not the same afferents. These findings need to be considered by investigators GROUP III RESPONSES TO CONTRACTION AND STRETCH using tendon stretch as a means to evoke the mechanical component of the exercise pressor reflex. 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