A natural history of efavirenz drug-induced liver injury.

BACKGROUND: Efavirenz (EFV), a non-nucleoside reverse transcriptase inhibitor, has been a component of first-line antiretroviral therapy (ART) in the South African HIV/AIDS programme since 2004. It is extensively used in ART programmes in other low- and middle-income countries. The natural history of the previously recognised EFV drug-induced liver injury (DILI) is not known. OBJECTIVES: To define and establish a causality assessment for EFV DILI and document its natural history by detailing a patient cohort. All relevant features characterising the patterns of clinical and histological injury, the duration of clinical and biochemical recovery and the associated mortality rate were documented. Factors associated with specific histological patterns of liver injury were analysed. METHODS: Patients were prospectively included after meeting causality and inclusion criteria for EFV DILI. Clinical, demographic and liver histological features (where possible) were documented from the time of presentation and throughout follow-up. Prednisone at 0.25 - 0.5 mg/kg was initiated at the discretion of the treating hepatologist. RESULTS: Fifty patients were prospectively included in the analysis. The median age was 34 (interquartile range (IQR) 29 - 39) years, males being older than females (p=0.014). Most (92%) were female, and 86% were of black African ethnicity. The median duration of ART at presentation was 6 months, with half of the women having initiated ART during pregnancy, at a median gestation of 24 (IQR 11 - 36) weeks. The median CD4 nadir at ART treatment initiation was 517 cells/µL, with no significant difference in CD4 nadir between those who were pregnant and those who were not (p=0.6). The median RUCAM (Roussel Uclaf Causality Assessment Method) score was 7, and among the 75% of patients who had liver biopsies, three histological patterns were identified: submassive necrosis (60%), nonspecific hepatitis (35%), and mixed cholestatic hepatitis (5%). On multivariate analysis, predictors for the development of submassive necrosis included younger age (<30 years; p=0.045), ART initiation in pregnancy (p=0.02), and a baseline CD4 count >350 cells/µL (p=0.018). For the nonspecific hepatitis group, pregnancy was also an associated factor (p=0.04). The mortality rate was 14%, with a median time from admission to death of 15 days. The median (IQR) time to initial hospital discharge was a lengthy 33 (24 - 52) days. Biochemical recovery was prolonged, necessitating a follow-up period of more than a year at an outpatient specialist clinic, with 86% of patients initiating a protease inhibitor-based ART regimen successfully. CONCLUSIONS: EFV DILI is a severe drug complication of ART with appreciable mortality and significant inpatient morbidity, requiring prolonged hospitalisation and follow-up.

The mechanics of setting up a COVID-19 response: Experiences of the COVID-19 epidemic from Groote Schuur Hospital, Cape Town, South Africa.

The SARS-CoV-2 pandemic has challenged the provision of healthcare in ways that are unprecedented in our lifetime. Planning for the sheer numbers expected during the surge has required public hospitals to de-escalate all non-essential clinical services to focus on COVID-19. Western Cape Province was the initial epicentre of the COVID-19 epidemic in South Africa (SA), and the Cape Town metro was its hardest-hit geographical region. We describe how we constructed our COVID-19 hospital-wide clinical service at Groote Schuur Hospital, the University of Cape Town's tertiary-level teaching hospital. By describing the barriers and enablers, we hope to provide guidance rather than a blueprint for hospitals elsewhere in SA and in low-resource countries that face similar challenges now or during subsequent waves.

Clinical management of COVID-19: Experiences of the COVID-19 epidemic from Groote Schuur Hospital, Cape Town, South Africa.

The SARS-CoV-2 pandemic has presented clinicians with an enormous challenge in managing a respiratory virus that is not only capable of causing severe pneumonia and acute respiratory distress syndrome, but also multisystem disease. The extraordinary pace of clinical research, and particularly the surge in adaptive trials of new and repurposed treatments, have provided rapid answers to questions of whether such treatments work, and has resulted in corticosteroids taking centre stage in the management of hospitalised patients requiring oxygen support. Some treatment modalities, such as the role of anticoagulation to prevent and treat potential thromboembolic complications, remain controversial, as does the use of high-level oxygen support, outside of an intensive care unit setting. In this paper, we describe the clinical management of COVID-19 patients admitted to Groote Schuur Hospital, a major tertiary level hospital at the epicentre of South Africa's SARS-CoV-2 epidemic during its first 4 months.

Alterations of myocardial dynamic stiffness implicating abnormal crossbridge function in human mitral regurgitation heart failure.

Mitral regurgitation (MR) causes ventricular dilation, a blunted myocardial force-frequency relation, and increased crossbridge force-time integral (FTI). The mechanism of FTI increase was investigated using sinusoidal length perturbation analysis to compare crossbridge function in skinned left ventricular (LV) epicardial muscle strips from 5 MR and 5 nonfailing (NF) control hearts. Myocardial dynamic stiffness was modeled as 3 parallel viscoelastic processes. Two processes characterize intermediate crossbridge cycle transitions, B (work producing) and C (work absorbing) with Q(10)s of 4 to 5. No significant differences in moduli or kinetic constants of these processes were observed between MR and NF. The third process, A, characterizes a nonenzymatic (Q(10)=0.9) work-absorbing viscoelasticity, whose modulus increases sigmoidally with [Ca(2+)]. Effects of temperature, crossbridge inhibition, or variation in [MgATP] support associating the calcium-dependent portion of A with the structural "backbone" of the myosin crossbridge. Extension of the conventional sinusoidal length perturbation analysis allowed using the A modulus to index the lifetime of the prerigor, AMADP crossbridge. This index was 75% greater in MR than in NF (P=0.02), suggesting a mechanism for the previously observed increase in crossbridge FTI. Notably, the A-process modulus was inversely correlated (r(2)=0.84, P=0.03) with in vivo LV ejection fraction in MR patients. The longer prerigor dwell time in MR may be clinically relevant not only for its potential role as a compensatory mechanism (increased economy of tension maintenance and increased resistance to ventricular dilation) but also for a potentially deleterious effect (reduced elastance and ejection fraction).

Myocardial cross-bridge kinetics in transition to failure in Dahl salt-sensitive rats.

The role of altered cross-bridge kinetics during the transition from cardiac hypertrophy to failure is poorly defined. We examined this in Dahl salt-sensitive (DS) rats, which develop hypertrophy and failure when fed a high-salt diet (HS). DS rats fed a low-salt diet were controls. Serial echocardiography disclosed compensated hypertrophy at 6 wk of HS, followed by progressive dilatation and impaired function. Mechanical properties of skinned left ventricular papillary muscle strips were analyzed at 6 wk of HS and then during failure (12 wk HS) by applying small amplitude (0.125%) length perturbations over a range of calcium concentrations. No differences in isometric tension-calcium relations or cross-bridge cycling kinetics or mechanical function were found at 6 wk. In contrast, 12 wk HS strips exhibited increased calcium sensitivity of isometric tension, decreased frequency of minimal dynamic stiffness, and a decreased range of frequencies over which cross bridges produce work and power. Thus the transition from hypertrophy to heart failure in DS rats is characterized by major changes in cross-bridge cycling kinetics and mechanical performance.

Protein osmotic pressure and the state of water in frog myoplasm.

We measured the osmotic pressure of diffusible myoplasmic proteins in frog (Rana temporaria) skeletal muscle fibers by using single Sephadex beads as osmometers and dialysis membranes as protein filters. The state of the myoplasmic water was probed by determining the osmotic coefficient of parvalbumin, a small, abundant diffusible protein distributed throughout the fluid myoplasm. Tiny sections of membrane (3.5- and 12-14-kDa cutoffs) were juxtaposed between the Sephadex beads and skinned semitendinosus muscle fibers under oil. After equilibration, the beads were removed and calibrated by comparing the diameter of each bead to its diameter measured in solutions containing 3-12% Dextran T500 (a long-chain polymer). The method was validated using 4% agarose cylinders loaded with bovine serum albumin (BSA) or parvalbumin. The measured osmotic pressures for 1.5 and 3.0 mM BSA were similar to those calculated by others. The mean osmotic pressure produced by the myoplasmic proteins was 9.7 mOsm (4 degrees C). The osmotic pressure attributable to parvalbumin was estimated to be 3.4 mOsm. The osmotic coefficient of the parvalbumin in fibers is approximately 3.7 mOsm mM(-1), i.e., roughly the same as obtained from parvalbumin-loaded agarose cylinders under comparable conditions, suggesting that the fluid interior of muscle resembles a simple salt solution as in a 4% agarose gel.

Changes in myofibrillar structure and function produced by N-terminal deletion of the regulatory light chain in Drosophila.

The similarity of amino acid sequence and motifs of the N-terminal extensions of certain class II myosin light chains, found throughout the animal kingdom, suggest a common functional role. One possible role of the N-terminal extension is to enhance oscillatory work and power production in striated muscles that normally operate in an oscillatory mode. We conducted small-angle X-ray diffraction experiments and small-length-perturbation analysis to examine the structural and functional consequences of deleting the N-terminal extension of the myosin regulatory light chain (RLC) in Drosophila flight muscle. The in vivo lattice spacing of dorsal longitudinal muscle (DLM) of flies lacking the RLC N-terminal extension (Dmlc2delta2-46) was approximately 1 nm less than that of wild type (48.56 +/- 0.02 nm). The myofilament lattice of detergent-treated, demembranated DLM swelled, with the DmlcdeltaA2-46 lattice expanding more than wild type and requiring roughly twice the concentration of Dextran T500 to restore its lattice to in vivo spacing (9-10% vs. 4% w/v). The calcium sensitivity and maximum amplitude of net oscillatory work near the in vivo lattice spacing was significantly lower in Dmlc2delta2-46 compared to wild type (pCa50 shifted by approximately one-third of a pCa unit; amplitude reduced by approximately one-half). These changes were in contrast to the lack of effect reported in a previous study carried out in the absence of Dextran T500. The results are consistent with the N-terminal extension interacting with actin to increase the probability that crossbridges form during stretch-activated oscillatory work and power production, especially at submaximal levels of calcium activation.

Role of the elastic protein projectin in stretch activation and work output of Drosophila flight muscles.

We examine how the stretch activation response of the Drosophila indirect flight muscles (IFM) is affected by the projectin mutation bentDominant. IFM from flies heterozygous for this mutation (bentD/+) produce approximately 85% full length projectin and approximately 15% truncated projectin lacking the kinase domain and more C-terminal sequences. Passive stiffness and power output of mutant fibers is similar to that of wild-type (+/+) fibers, but the amplitude of the stretch activation response (delayed tension rise) was significantly reduced. Measurement of actomyosin kinetics by sinusoidal analysis revealed that the apparent rate constant of the delayed tension rise (2 pi b) increased in proportion to the decrease in amplitude, accounting for the near wild-type levels of power output and nearly normal flight ability. These results suggest that projectin plays a crucial role in stretch activation, possibly through its protein kinase activity, by modulating crossbridge recruitment and kinetics.

In vivo x-ray diffraction of indirect flight muscle from Drosophila melanogaster.

Small-angle x-ray diffraction from isolated muscle preparations is commonly used to obtain time-resolved structural information during contraction. We extended this technique to the thoracic flight muscles of living fruit flies, at rest and during tethered flight. Precise measurements at 1-ms time resolution indicate that the myofilament lattice spacing does not change significantly during oscillatory contraction. This result is consistent with the notion that a net radial force maintains the thick filaments at an equilibrium interfilament spacing of approximately 56 nm throughout the contractile cycle. Transgenic flies with amino-acid substitutions in the conserved phosphorylation site of the myosin regulatory light chain (RLC) exhibit structural abnormalities that can explain their flight impairment. The I(20)/I(10) equatorial intensity ratio of the mutant fly is 35% less than that of wild type, supporting the hypothesis that myosin heads that lack phosphorylated RLC remain close to the thick filament backbone. This new experimental system facilitates investigation of the relation between molecular structure and muscle function in living organisms.

The effect of removing the N-terminal extension of the Drosophila myosin regulatory light chain upon flight ability and the contractile dynamics of indirect flight muscle.

The Drosophila myosin regulatory light chain (DMLC2) is homologous to MLC2s of vertebrate organisms, except for the presence of a unique 46-amino acid N-terminal extension. To study the role of the DMLC2 N-terminal extension in Drosophila flight muscle, we constructed a truncated form of the Dmlc2 gene lacking amino acids 2-46 (Dmlc2(Delta2-46)). The mutant gene was expressed in vivo, with no wild-type Dmlc2 gene expression, via P-element-mediated germline transformation. Expression of the truncated DMLC2 rescues the recessive lethality and dominant flightless phenotype of the Dmlc2 null, with no discernible effect on indirect flight muscle (IFM) sarcomere assembly. Homozygous Dmlc2(Delta2-46) flies have reduced IFM dynamic stiffness and elastic modulus at the frequency of maximum power output. The viscous modulus, a measure of the fly's ability to perform oscillatory work, was not significantly affected in Dmlc2(Delta2-46) IFM. In vivo flight performance measurements of Dmlc2(Delta2-46) flies using a visual closed-loop flight arena show deficits in maximum metabolic power (P(*)(CO(2))), mechanical power (P(*)(mech)), and flight force. However, mutant flies were capable of generating flight force levels comparable to body weight, thus enabling them to fly, albeit with diminished performance. The reduction in elastic modulus in Dmlc2(Delta2-46) skinned fibers is consistent with the N-terminal extension being a link between the thick and thin filaments that is parallel to the cross-bridges. Removal of this parallel link causes an unfavorable shift in the resonant properties of the flight system, thus leading to attenuated flight performance.

Morphology and transverse stiffness of Drosophila myofibrils measured by atomic force microscopy.

Atomic force microscopy was used to investigate the surface morphology and transverse stiffness of myofibrils from Drosophila indirect flight muscle exposed to different physiologic solutions. I- and A-bands were clearly observed, and thick filaments were resolved along the periphery of the myofibril. Interfilament spacings correlated well with estimates from previous x-ray diffraction studies. Transverse stiffness was measured by using a blunt tip to indent a small section of the myofibrillar surface in the region of myofilament overlap. At 10 nm indention, the effective transverse stiffness (K( perpendicular)) of myofibrils in rigor solution (ATP-free, pCa 4.5) was 10.3 +/- 5.0 pN nm(-1) (mean +/- SEM, n = 8); in activating solution (pCa 4.5), 5.9 +/- 3.1 pN nm(-1); and in relaxing solution (pCa 8), 4.4 +/- 2.0 pN nm(-1). The apparent transverse Young's modulus (E( perpendicular)) was 94 +/- 41 kPa in the rigor state and 40 +/- 17 kPa in the relaxed state. The value of E( perpendicular) for calcium-activated myofibrils (55 +/- 29 kPa) was approximately a tenth that of Young's modulus in the longitudinal direction, a difference that at least partly reflects the transverse flexibility of the myosin molecule.

Parvalbumin concentration and diffusion coefficient in frog myoplasm.

The concentrations and diffusivity of two isoforms of parvalbumin, IVa and IVb, were measured using quantitative SDS PAGE in single fibers from semitendinosus muscles of the frog Rana temporaria. The concentrations of IVa and IVb were 2.9 +/- 0.3 (SEM) and 4.5 +/- 0.5 g l-1 total fiber volume, respectively. The total concentration of parvalbumin (7.4 +/- 0.8 g l-1 total fiber) corresponds to a cytosolic concentration of 0.9 +/- 0.1 mmol l-1 myoplasmic water. Estimates for the transverse and longitudinal diffusion coefficients for parvalbumin at 4 degrees C were obtained in two ways: (1) by diffusion of parvalbumin out of skinned fibers into droplets of relaxing solution, and (2) by diffusion of parvalbumin between two juxtaposed skinned fibers under oil. The transverse diffusion coefficient obtained using the droplet method was significantly lower than that obtained using juxtaposed fibers, but the longitudinal diffusion coefficients obtained from both methods were similar. The juxtaposed fiber method more accurately approximates parvalbumin diffusion in undisturbed myoplasm because no artificial solutions were used and, upon fiber-to-fiber contact, a potentially confounding oil barrier at the interface rapidly disperses. The juxtaposed fiber method yielded values for transverse (4.27 +/- 0.87 x 10(-7) cm2 s-1) and longitudinal (3.20 +/- 0.74 x 10(-7) cm2 s-1) diffusion coefficients that were not significantly different, suggesting that diffusion of parvalbumin in myoplasm is essentially isotropic. The average diffusion coefficient of frog parvalbumin in myoplasm (3.74 +/- 0.81 x 10(-7) cm2 s-1; 4 degrees C) is approximately a third of that estimated for frog parvalbumin diffusing in bulk water into and out of 3% agarose cylinders (10.6 x 10(-7) cm2 s-1; 4 degrees C). The reduced translational mobility of parvalbumin in myoplasm reflects an elevated effective viscosity due to tortuosity and viscous drag imposed by the fixed proteins of the cytomatrix and the numerous diffusible particles of the cytosol.

Altered crossbridge kinetics in the alphaMHC403/+ mouse model of familial hypertrophic cardiomyopathy.

A mutation in the cardiac beta-myosin heavy chain, Arg403Gln (R403Q), causes a severe form of familial hypertrophic cardiomyopathy (FHC) in humans. We used small-amplitude (0.25%) length-perturbation analysis to examine the mechanical properties of skinned left ventricular papillary muscle strips from mouse hearts bearing the R403Q mutation in the alpha-myosin heavy chain (alphaMHC403/+). Myofibrillar disarray with variable penetrance occurred in the left ventricular free wall of the alphaMHC403/+ hearts. In resting strips (pCa 8), dynamic stiffness was approximately 40% greater than in wild-type strips, consistent with elevated diastolic stiffness reported for murine hearts with FHC. At pCa 6 (submaximal activation), strip isometric tension was approximately 3 times higher than for wild-type strips, whereas at pCa 5 (maximal activation), tension was marginally lower. At submaximal calcium activation the characteristic frequencies of the work-producing (b) and work-absorbing (c) steps of the crossbridge were less in alphaMHC403/+ strips than in wild-type strips (b=11+/-1 versus 15+/-1 Hz; c= 58+/-3 versus 66+/-3 Hz; 27 degrees C). At maximal calcium activation, strip oscillatory power was reduced (0. 53+/-0.25 versus 1.03+/-0.18 mW/mm3; 27 degrees C), which is partly attributable to the reduced frequency b, at which crossbridge work is maximum. The results are consistent with the hypothesis that the R403Q mutation reduces the strong binding affinity of myosin for actin. Myosin heads may accumulate in a preforce state that promotes cooperative activation of the thin filament at submaximal calcium but blunts maximal tension and oscillatory power output at maximal calcium. The calcium-dependent effect of the mutation (whether facilitating or debilitating), together with a variable degree of fibrosis and myofibrillar disorder, may contribute to the diversity of clinical symptoms observed in murine FHC.

The Drosophila projectin mutant, bentD, has reduced stretch activation and altered indirect flight muscle kinetics.

Projectin is a ca. 900 kDa protein that is a member of the titin protein superfamily. In skeletal muscle titins are involved in the longitudinal reinforcement of the sarcomere by connecting the Z-band to the M-line. In insect indirect flight muscle (IFM), projectin is believed to form the connecting filaments that link the Z-band to the thick filaments and is responsible for the high relaxed stiffness found in this muscle type. The Drosophila mutant bentD (btD) has been shown to have a breakpoint close to the carboxy-terminal kinase domain of the projectin sequence. Homozygotes for btD are embryonic lethal but heterozygotes (btD/+) are viable. Here we show that btD/+ flies have normal flight ability and a slightly elevated wing beat frequency (btD/+ 223+/-13 Hz; +/+ 203+/-5 Hz, mean +/- SD; P < 0.01). Electron microscopy of btD/+ IFM show normal ultrastructure but skinned fiber mechanics show reduced stretch activation and oscillatory work. Although btD/+ IFM power output was at wild-type levels, maximum power was achieved at a higher frequency of applied length perturbation (btD/+ 151+/-6 Hz; +/+ 102+/-14 Hz; P < 0.01). Results were interpreted in the context of a viscoelastic model of the sarcomere and indicate altered cross-bridge kinetics of the power-producing step. These results show that the btD mutation reduces oscillatory work in a way consistent with the proposed role of the connecting filaments in the stretch activation response of IFM.

Mechanoenergetic alterations during the transition from cardiac hypertrophy to failure in Dahl salt-sensitive rats.

BACKGROUND: The time course and mechanisms of altered mechanoenergetics and depressed cross-bridge cycling in hypertrophied and failing myocardium are uncertain. METHODS AND RESULTS: We studied mechanoenergetics in Dahl salt-sensitive (DS) rats fed high-salt diet (HS) for 6 (HS-6) and 12 (HS-12) weeks to produce compensated hypertrophy and failure. The slope of the end-systolic pressure-volume relation (E'max) was similar in HS-6 and low-salt controls (LS-6), but reduced in HS-12 compared with controls (LS-12). Efficiency [1/slope of oxygen consumption (&f1;O2)-pressure-volume area (PVA) relation] was similar in HS-6 and LS-6 but higher in HS-12 versus LS-12 (59+/-16% versus 44+/-7%, P<0.05). Economy [1/slope of the force-time integral (FTI)-&f1;O2 relation] was similar in HS-6 and LS-6 but higher in HS-12 versus LS-12 (218+/-123 versus 74+/-39x10(3) g. s. mL O2-1. g; P<0.05). Compared with controls, myofibrillar ATPase activity was reduced by 24% in HS-6 and 44% in HS-12. V3 Isomyosin was increased in HS-6 (40+/-12% versus 9+/-8%; P<0.05) and further increased in HS-12 (76+/-10% versus 22+/-18%; P<0.05). Hypothyroid LS-12 rats had 100% V3 isomyosin, yet efficiency, economy, and ATPase values were intermediate between LS-12 and HS-12. HS-12 rats demonstrated increased troponin T3 isoform (17+/-2 versus 23+/-2%, P<0.05). There were no changes in troponin I or tropomyosin isoforms. However, the proportion of phosphorylated troponin T was reduced in HS-12 versus LS-12 hearts (P<.001). CONCLUSIONS: In DS rats, the transition to failure is associated with depressed E'max and increased efficiency and economy. These findings are linked to myofibrillar ATPase activity and suggest that mechanisms other than isomyosin switching are important determinants of ventricular energetics. A troponin T isoform switch is one potential mechanism.

A genetic deficiency that spans the flightin gene of Drosophila melanogaster affects the ultrastructure and function of the flight muscles.

We have developed a reverse-genetic approach to study the function of flightin, a unique protein of the flight muscle myofibril of Drosophila melanogaster. We describe the generation and characterization of Df(3L)fln1, a lethal genetic deficiency in the 76BE region of the third chromosome which deletes several genes, including the gene for flightin. We show that heterozygous flies harboring the Df(3L)fln1 mutation exhibit both impaired flight and ultrastructural defects in their flight muscle myofibrils. We found that the mutation does not interfere with assembly of the myofibril but leads to disorganization of peripheral myofilaments in adult myofibrils. Most myofibrils, nevertheless, retain an intact core that represents approximately 80 % of the normal lattice diameter. Mechanical analysis of single skinned flight muscle fibers demonstrates that the mutation has no significant effect on net power output but increases the frequency at which maximum power is delivered to the wings, potentially reducing the overall performance of the flight system. The results suggest that flightin is an indispensable part of the flight muscle contractile mechanism.

Functional significance of cardiac myosin essential light chain isoform switching in transgenic mice.

The different functions of the ventricular- and atrial-specific essential myosin light chains are unknown. Using transgenesis, cardiac-specific overexpression of proteins can be accomplished. The transgenic paradigm is more useful than originally expected, in that the mammalian heart rigorously controls sarcomeric protein stoichiometries. In a clinical subpopulation suffering from heart disease caused by congenital malformations of the outflow tract, an ELC1v-->ELC1a isoform shift correlated with increases in cross-bridge cycling kinetics as measured in skinned fibers derived from the diseased muscle. We have used transgenesis to replace the ventricular isoform of the essential myosin light chain with the atrial isoform. The ELC1v--> ELC1a shift in the ventricle resulted in similar functional alterations. Unloaded velocities as measured by the ability of the myosin to translocate actin filaments in the in vitro motility assay were significantly increased as a result of the isoform substitution. Unloaded shortening velocity was also increased in skinned muscle fibers, and at the whole organ level, both contractility and relaxation were significantly increased. This increase in cardiac function occurred in the absence of a hypertrophic response. Thus, ELC1a expression in the ventricle appears to be advantageous to the heart, resulting in increased cardiac function.

Work production and work absorption in muscle strips from vertebrate cardiac and insect flight muscle fibers.

Stretch activation, which underlies the ability of all striated muscles to do oscillatory work, is a prominent feature of both insect flight and vertebrate cardiac muscle. We have examined and compared work-producing and work-absorbing processes in skinned fibers of Drosophila flight muscle, mouse papillary muscle, and human ventricular strips. Using small amplitude sinusoidal length perturbation analysis, we distinguished viscoelastic properties attributable to crossbridge processes from those attributable to other structures of the sarcomere. Work-producing and work-absorbing processes were identified in Ca(2+)-activated fibers by deconvolving complex stiffness data. An 'active' work-producing process ("B"), attributed to crossbridge action, was identified, as were two work-absorbing processes, one attributable to crossbridge action ("C") and the other primarily to viscoelastic properties of parallel passive structures ("A"). At maximal Ca(2+)-activation (pCa 5, 27 degrees C), maximum net power output (processes A, B and C combined) occurs at a frequency of: 1.3 +/- 0.1 Hz for human, 10.9 +/- 2.2 Hz for mouse, and 226 +/- 9 Hz for fly, comparable to the resting heart rate of the human (1 Hz, 37 degrees C) and mouse (10 Hz, 37 degrees C) and to the wing beat frequency of the fruit fly (200 Hz, 22 degrees C). Process B maximal work production per myosin head is 7-11 x 10(-21) J per perturbation cycle, equivalent to approximately 2 kT of energy. Process C maximal work absorption is about the same magnitude. The equivalence suggests the possibility that a thermal ratchet type mechanism operates during small amplitude length perturbations. We speculate that there may be a survival advantage in having a mechanical energy dissipater (i.e., the C process) at work in muscles if they can be injuriously stretched by the system in which they operate.