A low-cost, multi-channel, EMG signal processing amplifier.

There is a growing need in studies of movement control to expand the number of muscles from which EMG activity is recorded during performance of motor tasks. Optimal viewing and analysis of this EMG activity requires signal processing which provides adjustable gain and baseline offset as wells as selectable AC coupling, rectification and filtering. This paper presents a low-cost circuit that combines two channels of EMG signal processing capability in one module.

Diaphragm shortening and intrathoracic pressure during hypercapnia in rats.

The purpose of this study was to determine the relationship between intrathoracic pressure (delta ITP) and diaphragm shortening (DS) during the development of diaphragm fatigue. Fatigue of the diaphragm was produced by having rats breath 15% CO2 in O2. Diaphragm shortening increased significantly to 178% of control during the first 5 min of hypercapnia and then decreased to 86% of control at approximately 80 min. Twenty minutes after terminating hypercapnia, DS increased to 115% of the prehypercapnic value. delta ITP increased to 199% of control following 5 min of hypercapnia and continued to increase, reaching 267% of control at the end of the hypercapnic period. Twenty minutes later, delta ITP was 147% of control. These results illustrate that during increased respiratory work, DS can decrease while intrathoracic pressure remains increased. These findings suggest that intrathoracic pressure may not always reflect the contractile status of the diaphragm. These findings are consistent with other studies indicating that as the diaphragm fatigues, accessory respiratory muscle activity increases to maintain delta ITP.

Intracellular magnesium affects I(K) in single frog atrial cells.

Voltage-clamp experiments on single frog (Rana pipiens) atrial cells using whole cell recording techniques revealed that the addition of MgCl2 to the 150 mM KCl patch pipette solution influenced the voltage- and time-dependent potassium current (IK). After rupture of the membrane patch under the tip of the pipette, IK increased with time when the pipette solution was magnesium free, but decreased slightly when the solution contained 1.5 mM MgCl2. More dramatic decreases in IK occurred when the solution contained 3.0 or 10 mM MgCl2. In addition to suppressing the magnitude of IK, the activation rate of this current was enhanced by 10 mM MgCl2 but was not affected by 1.5 or 3 mM MgCl2. Other chloride salts containing mono-, di-, or trivalent cations were used to demonstrate that the effects of MgCl2 on IK were not related to alterations in ionic strength, osmolality, or chloride concentration produced by adding MgCl2 to the pipette solution. Our results suggest that changes in the intracellular magnesium concentration influence IK as the pipette solution exchanges with the intracellular fluid.

Voltage-tension relations in single frog atrial cardiac cells.

Voltage-clamp experiments were performed on isolated single frog (Rana catesbeiana or Rana pipiens) atrial cells to determine the voltage-contraction relations of the single cardiac cell. The contractile responses of the single cell associated with long duration (3 second) depolarizing steps consisted of a rise to peak (phasic) followed by a decay to a sustained contraction (tonic). These phasic-tonic type contractile responses could be obtained under conditions where membrane potential was well controlled along the entire length of the cell. Thus, the data obtained on the single cell indicate that the phasic-tonic contractile response is the characteristic contractile response of frog atrial tissue. The voltage dependence of the extent of relaxation to the tonic component following the peak of the contraction was affected dramatically by the intracellular sodium concentration. This result indicates that both the relaxation following peak contraction as well as the tonic contraction are related to calcium control via the sodium-calcium exchanger. The data also indicate that calcium entry via the inward calcium current is required for the contractile response to have a phasic component. These data indicate that calcium entry via the inward calcium current followed by the sodium-calcium exchanger first reducing and then maintaining the intracellular calcium level produces the characteristic phasic-tonic contractile response.

Implementation of a movement paradigm using the Commodore 64 microcomputer.

Implementation of an alternating movement paradigm for monkeys was achieved using an inexpensive but versatile microcomputer, the Commodore 64. During task performance, the computer monitors one of three user selectable input signals (e.g. joint position) and continuously displays this signal as a moving cursor on a video monitor along with a user positioned target box. Other user defined parameters include in-target holding time and reinforcement ratio. The system also provides two sound cues to signal entry of the cursor into the target box and successful completion of a trial. Extensive use is made of the computer's intrinsic hardware features for implementation of movement paradigm functions. Use of external components is limited to digitizing and interface hardware. A two part software package consisting of a BASIC and a machine language program performs all task and hardware related functions. Acquisition and display of analog input signals, display of target positions, and delivery of auditory cues and applesauce rewards are all controlled by the machine language program. All user defined parameters are specified from the BASIC menu program. The specific programs described in this paper should be applicable to the control of tasks requiring alternation of a behavioral parameter between two target zones.

Depression of contractility following stretches and releases applied during contraction to single frog atrial cardiac cells.

The effects of stretches and releases on the contractile performance of isolated single frog atrial cells (Rana catesbeiana) were investigated. A stretch or release was imposed on the cell--either during a contraction (test) or before the onset of contraction (control)--and the contractile performance (length, velocity and force) of the test contraction was compared with that of the control contraction to determine whether the stretch or release imposed on the contracting cell altered the contractility of the cell. We found that the velocity of cell (and sarcomere) shortening for the remainder of the test contraction following either a stretch or release was markedly less than that occurring at the same time in the control contraction. This decreased velocity occurred even though the force in the test contraction was less than that in the control contraction and the sarcomere length was longer in the test contraction than in the control contraction. These results indicate that after a stretch or release imposed on the contracting cell, the force-velocity relationship at any given length and time is depressed than had the stretch or release not been imposed on the contracting cell. Thus, stretches and releases applied to the contracting single cardiac cell either produce a long-term depression in the contractility of the cell, or that the contractility at any given time and sarcomere length depends markedly on the history of the contraction.

Arterial baroreceptor reflex control of heart rate in two species of turtle.

Attempts were made to stimulate an arterial baroreceptor reflex in anesthetized and conscious pond turtles. In turtles anesthetized with either alpha-chloralose or pentobarbital, occlusion of the ascending or descending aortas produced no reflex heart rate (HR) changes. In pentobarbital-anesthetized turtles, direct electrical or mechanical stimulation of potential baroreceptor sites along the central aortic arches and carotid arteries produced no significant changes in either HR or blood pressure (BP). Occlusion of the common carotid arteries also produced no HR or BP changes. Intravenously administered nitroglycerin lowered BP but caused no reflex tachycardia in anesthetized turtles. Phenylephrine and angiotensin elevated BP in the anesthetized turtle but caused no reflex bradycardia. In conscious turtles phenylephrine increased BP and nitroglycerin decreased BP. Neither response produced a statistically significant HR change, although HR tended to decrease transiently with phenylephrine and increase with nitroglycerin. These HR changes were abolished by atropine. Rapid intra-arterial infusion of 6% dextran transiently raised BP but caused no reflex bradycardia. These experiments suggest that cardiovascular regulation in the turtle is accomplished without a major contribution from arterial baroreceptor reflexes.

Effect of external force on relaxation kinetics in single frog atrial cardiac cells.

The effects of external force on relaxation kinetics were investigated in isolated single frog (Rana catesbeiana) atrial cells. We found that force decay occurred at a maximum and constant rate for a significant portion of auxotonic relaxation, and this rate was linearly related to the peak force developed during auxotonic contraction. The slope of the linear relationship between the maximum rate of auxotonic force decay and peak auxotonic force was not affected by changes in the level of contractile activation produced by activating the cell with different stimulus durations. The rate of force change during auxotonic contraction and relaxation in the isolated cell is directly related to the average sarcomere velocity within the cell. Thus, the results indicate that during auxotonic relaxation the velocity of sarcomere extension is directly related to the peak auxotonic force, and sarcomere extension, during relaxation, is therefore affected by external force. The direct effect of external force on relaxation kinetics was confirmed by the observation that force changes imposed on the cell during relaxation immediately altered the velocity of the extending cell from any given length. However, data are also presented which demonstrate that rapid sarcomere extension occurs during relaxation under conditions where external forces are negligible. Thus, rapid sarcomere extension during relaxation does not require large external forces, and internal forces must play a role in sarcomere extension during relaxation. An explanation is given for these apparently contradictory results.

Circulatory mechanoreceptors in the pond turtle Pseudemys scripta.

This study was undertaken to characterize cardiovascular receptors in the turtle, Pseudemys scripta, with particular attention being given to neural activity changes associated with alterations in blood pressure. Vagal afferent nerve traffic, synchronous with heart contractions, was recorded in anesthetized artificially ventilated turtles. Action potentials, from receptors that fired regularly during each heart cycle, occurred during ventricular systole. Mechanical probing and vascular occlusion indicated that these receptors were located in the proximal common pulmonary artery including the bulbus cordis region. Bolus injections of saline into the ventricle or the common pulmonary artery caused immediate but transient increases in cardiac synchronous traffic. Prolonged elevation of arterial and ventricular blood pressure, by either saline injection or arterial occlusion, caused increases in receptor discharge of the same duration as the pressure increases. Although these receptors could participate in the regulation of the systemic and the pulmonary circulation, the physiological role for them is presently unknown.

Effect of initial sarcomere length on sarcomere kinetics and force development in single frog atrial cardiac cells.

We studied sarcomere performance in single isolated intact frog atrial cells using techniques that allow direct measurement of sarcomere length and force. The purpose of this investigation was to determine whether length-dependent alterations in contractile activation occur in the single isolated cardiac cell. This was accomplished by determining the effect of initial sarcomere length on the time course of sarcomere shortening and force development during auxotonic twitch contractions. The results presented in this paper demonstrate that the velocity of sarcomere shortening, the rate of force development, and the magnitude of force development during auxotonic twitch contractions all increase as initial sarcomere length increases over the range of about 2 micrometers to greater than 3 micrometers. These results indicate that the level of contractile activation increases as initial sarcomere length increases. Also, results are presented that indicate that the rate of increase of contractile activation during a twitch contraction also increases as initial sarcomere length increases. These length-dependent effects on contractile activation in conjunction with the slow time course of contractile activation cause the force-velocity-length relationship to be time-dependent: i.e., the velocity of sarcomere shortening at a given sarcomere length and load depends on the time during the contraction when the sarcomere reaches that length. The results suggest that length-dependent alterations in contractile activation may play a major role in the improved contractile performance that accompanies an increase in initial sarcomere length in cardiac muscle.

Characteristics of sarcomere shortening in single frog atrial cardiac cells during lightly loaded contractions.

We studied sarcomere performance in single isolated intact cardiac cells using techniques that allow direct measurement of sarcomere length and force. This investigation dealt primarily with sarcomere performance during twitch contractions under lightly loaded conditions. In such contractions, there was a significant portion of the contraction in which sarcomere shortening occurred at constant velocity over a significant range of sarcomere lengths. The constant velocity phase of shortening was followed by a phase of shortening in which sarcomere velocity decreased markedly. Both the velocity and extent of sarcomere shortening depended on the stimulus parameters used to excite the cell. With threshold stimulation, sarcomere velocities during the constant velocity phase of shortening ranged from 1 to 5.5 micron/sec in different cells and significant slowing of sarcomere shortening began at sarcomere lengths of 1.8-2.0 micron. In contrast, when cells were stimulated with a long duration stimulus (200 msec) of large current strength, sarcomere velocities during the constant velocity phase ranged from 6 to 12 micron/sec, and significant slowing did not occur until a sarcomere length of about 1.6 micron was reached. The threshold stimulus strength-stimulus duration relationship was determined on the single cell, and it was found to be of the type expected for a cell having an intact excitable membrane capable of generating an action potential when depolarized to a fixed voltage threshold. The data presented in this paper give direct evidence that the lightly loaded cardiac sarcomere has a velocity of shortening which depends on the level of contractile activation but is independent of sarcomere length at sarcomere lengths greater than about 1.6 micron.

Evidence that the velocity of sarcomere shortening in single frog atrial cardiac cells is load dependent.

Recent experiments using laser diffraction techniques to determine the time course and extent of sarcomere shortening in thin bundles of cardial tissue have given results which suggest that the velocity of sarcomere shortening in cardiac muscle is independent of the developed force (Nassar et al., 1974; Krueger and Pollack, 1975). However, the anatomical complexity of the intact tissue precludes a definite interpretation of the data, since the exact relationship between the force being borne by the total tissue to the force being borne by any observed group of sarcomeres is uncertain. The single frog atrial cell provides a simple cardiac preparation in which the relationship between sarcomere velocity and sarcomere force is well defined, since these cells are only 1-2 myofibrils wide. The purpose of the present investigation was to determine if sarcomere velocity in the single frog atrial cell is dependent on force by measuring the time course of sarcomere shortening in single cells under conditions in which the cell developed markedly different forces. The results presented in this paper give direct evidence that the velocity of sarcomere shortening in the single cardiac cell depends on the force being developed by the sarcomeres. Thus, cardiac sarcomeres have a type of force-velocity relationship, although the exact nature of this relationship could not be determined in these experiments.

Sarcomere length-resting tension relation in single frog atrial cardiac cells.

It generally has been thought that the relatively high resting tension characteristic of cardiac tissue resides in structures (collagen, elastin) external to the individual cardiac cells, but the evidence to support this conclusion has been indirect, since the resting tension of intact single cardiac cells has not been determined previously. The purpose of the present investigation was to determine the resting tension (stress)-sarcomere length relationships of single intact frog atrial cells. For tension determinations, a single cell was attached between two poly-L-lysine coated glass beams; one beam served as a compliant calibrated cantilevered force beam, and length changes were imposed on the cell by movement of the other beam. Coventional bright-field light microscope techniques were used to view the cell, the sarcomere pattern within the cell, and the position of the force beam. The resting tension of the intact cell increased from a value of about 10 nN at a sarcomere length of 2.35 microns to a value of about 130 nN at a sarcomere length of 3.45 microns. Lagrangian and Eulerian resting stress-sarcomere length relationships were computed from the resting tension-sarcomere length relationships. The Lagrangian stress increased from a value of about 0.6 mN/mm2 at a sarcomere length of 2.35 microns to a value of about 7 mN/mm2 at a sarcomere length of 3.45 microns. These values of stress are about 8- to 30-fold less than those previously reported for intact frog atrial tissue and indicate that the resting tension of intact frog atrial preparations resides primarily in structures external to the individual cardiac cell.

Chemically mediated cell-to-cell contractile activation in isolated frog atrial cardiac cells.

A dying single frog atrial cardiac cell liberates an unknown substance which diffuses away from the dying cell and activates contractile activity in other isolated intact single cardiac cells within the vicinity of the dying cell.

Limitations of the double sucrose gap voltage clamp technique in tension-voltage determinations on frog atrial muscle.

The purpose of this study was to evaluate the limitations of the double sucrose gap voltage clamp technique in the determination of tension-voltage relationships for frog atrial muscle. Tension-voltage relationships were determined under two conditions. In one case we determined both the tension response and slow inward current associated with an apparent step depolarization (step-clamp) as a function of the magnitude of the step depolarization. In the second case, an action potential was elicited, the voltage clamp was applied early during the plateau phase of the action potential, and the tension response was determined as a function of the clamp potential (action potential-clamp). Under both step-clamp and action potential-clamp conditions, the waveform of the tension response rose to a peak value (Tp) and then decayed with time to a tension that was maintained for the duration of the depolarization. The Tp-clamp potential relationships obtained under step-clamp and action potential-clamp conditions were similar. Microelectrode measurements of transmembrane potential of cells in the "voltage-clamped" region of the preparation demonstrated the lack of temporal and spatial voltage control under both step-clamp and action potential-clamp conditions, and also demonstrated that acquisition of spatial voltage control occurred at about the same time that the tension response reached its peak value. These data indicate that this voltage clamp technique does not allow an accurate determination of the so-called phasic tension-voltage relationship in frog atrial muscle because of a lack of temporal and spatial control of voltage during the rising phase of the tension response.

Preparation of isolated single cardiac cells from adult frog atrial tissue.

Isolated cardiac cells from bullfrog atrial tissue can be readily prepared by digestion of intact fragments of atrial tissue with trypsin and collagenase. These isolated cells have dimensions of about 5 mum in width and range in length from 300 mum to over 500 mum. Such isolated cells may prove useful for the investigation of contractile activity of cardiac muscle at the single cell level and at the sarcomere level within the single cell.

An assessment of the double sucrose-gap voltage clamp technique as applied to frog atrial muscle.

The homogeneity of voltage clamp control in small bundles of frog atrial tissue under double sucrose-gap voltage clamp conditions was assessed by intracellular microelectrode potential measurements from cells in the test node region. The microelectrode potential measurements demonstrated that (1) good voltage control of the impaled cell existed in the absence of the excitatory inward currents (e.g., during small depolarizing clamp pulses of 10-15 mV), (2) voltage control of the impaled cell was lost during either the fast or slow excitatory inward currents, and (3) voltage control of the impaled cell was regained following the inward excitatory currents. Under nonvoltage clamp conditions the transgap recorded action potential had a magnitude and waveform similar to the intracellular microelectrode recorded action potentials from cells in the test node. Transgap impedance measured with a sine-wave voltage of 1,000 Hz was about 63% of that measured either by a sine-wave voltage of 10 Hz or by an action potential method used to determine the longitudinal resistance through the sucrose-gap region. The action potential data in conjunction with the impedance data indicate that the extracellular resistance (R(s)) through the sucrose gap is very large with respect to the longitudinal intracellular resistance (R(i)); the frequency dependence of the transgap impedance suggests that at least part of the intracellular resistance is paralleled by a capacitance. The severe loss of spatial voltage control during the excitatory inward current raises serious doubts concerning the use of the double sucrose-gap technique to voltage clamp frog atrial muscle.