Determination of the force necessary for the propagation of tears in ostrich and calf pericardium

The durability of prosthetic heart valve leaflets made of biological materials is limited. A tear in the biomaterial accelerates their early failure, but microtearing of the collagen fibers may be responsible for their medium-term failure. We studied the force necessary to propagate tearing in two biomaterials: ostrich and calf pericardium. One hundred twenty samples of each tissue were tested in an Elmendorf pendulum capable of measuring the force required to tear a tissue in which a predefined slit had been made. The forces required to produce tears, ranging between 2.5 and 0.25 cm in length, were determined. For ostrich pericardium, this force ranged between 67.67 and 4.80 newton, while that required to tear the same lengths of calf pericardium ranged between 70.67 and 4.70 newton. The function that relates the tearing force to the length of the tear was expressed as follows: y = 20.62x + 1.77x(2) (R(2) = 0.923) for ostrich pericardium and y = 45.57x - 7.21x(2) (R(2) = 0.936) for calf pericardium, where y is the force in newton and x is the length in centimeter. Calf pericardium was found to have a greater resistance to tearing. However, these results should be interpreted with caution owing to the fact that the thickness of the majority of the samples of ostrich pericardium was significantly less than that of calf pericardium. A more careful selection and utilization of adult ostrich pericardium would probably improve these results.     

Comparison of the mechanical behaviors of biological tissues subjected to uniaxial tensile testing: pig,calf and ostrich pericardium sutured with Gore-Tex

The purpose of this study was to compare the mechanical behavior of calf pericardium, pig pericardium and ostrich pericardium when subjected to tensile testing. Tensile stress was applied to 108 tissue samples, 36 of each type of tissue, until rupture. Groups of three adjacent strips measuring 12 x 2 cm(2) were cut longitudinally. Each group consisted of an unsutured center sample, or control, and the two contiguous samples, that on the right sutured with Gore-Tex at a 90 degrees angle with respect to the longitudinal axis and that on the left sewn with the same suture material at 45 degrees angle. The sutured samples showed a statistically significant loss of resistance (p<0.001) when compared with the corresponding unsutured tissue. The mean stresses at rupture for sutured ostrich pericardium were 21.81 and 20.81 MPa in the samples sewn at 45 degrees and 90 degrees, respectively, higher than those corresponding to unsutured calf and pig pericardium, 14.0 and 11.49 MPa, respectively, at rupture.The analysis of the stress/strain curve shows a smaller difference between sutured and unsutured ostrich pericardium than those observed in the other two biomaterials. These results demonstrate that, in addition to its greater resistance, ostrich pericardium also presents a less pronounced interaction with the suture material. Its capacity to absorb the shearing stress produced by the suture is greater. This report also confirms that the method of selection using paired samples ensures their homogeneity and makes it possible to predict the behavior of a sample by determining that of the other half of the pair.     

Resistance to tearing of calf and ostrich pericardium: Influence of the type of suture material and the direction of the suture line

The tearing of the valve leaflet of a cardiac bioprosthesis can cause early failure of this device, which is employed to replace a diseased native valve. This report involves the study of the behavior of 312 tissue samples (152 of calf pericardium and 160 of ostrich pericardium) treated with glutaraldehyde and subsequently subjected to tear testing. The samples were cut in the two principal directions: longitudinally, or root to apex, and transversely. They included a series of control samples that were left unsutured, and the remaining samples were repaired with the use of two different suture techniques: a running suture in the direction of the load and a telescoping suture perpendicular to the load. Four commercially available suture materials were employed: Pronova, nylon, Gore-Tex, or silk. The unsutured control samples of both types of pericardium exhibited a similar anisotropic behavior in the tear test. The mean resistance to tearing of the calf pericardium was 24.29 kN m in samples cut longitudinally and 34.78 kN m in those cut transversely (p =.03); the values were 28.08 kN m and 37.12 kN m (p =.002), respectively, in ostrich pericardium. The series repaired with the telescoping suture always exhibited greater resistance to tearing, with values that ranged between 44.34 and 64.27 kN for the samples of calf pericardium and from 41.65 to 47.65 kN for those obtained from ostrich. These assays confirm the anisotropic behavior of calf and ostrich pericardium treated with glutaraldehyde when subjected to tear testing, as well as the loss of this behavior in ostrich pericardium after suturing. Suturing techniques, such as the telescoping model, that provide a greater resistance to tearing should be studied for use in the design of the valve leaflets of cardiac bioprostheses made of biological materials.     

Biochemical and mechanical behavior of ostrich pericardium as a new biomaterial

We have performed a comparative analysis of glutaraldehyde-preserved ostrich pericardium, as a novel biomaterial, with bovine pericardium. The biochemical characteristics (histology, water content, amino acid composition, and collagen and elastin contents), mechanical properties, and in vivo calcification in a subcutaneous rat model were examined. Ostrich pericardium is slightly thinner and shows a higher water content (70+/-2% vs. 62+/-2%) than bovine pericardium. Additionally, ostrich pericardium presents 1.6-fold lower elastin content and a lower percentage of collagen in reference to the total protein content (68+/-2% vs. 76+/-2%). However, ostrich pericardium shows better mechanical properties, with higher tensile stress at rupture (32.4+/-7.5 vs. 11.5+/-4.6) than calf pericardium. In vivo calcification studies in a rat subcutaneous model show that ostrich pericardium is significantly less calcified than bovine pericardium (23.95+/-13.30 vs. 100.10+/-37.36 mg/g tissue) after 60 days of implantation. In conclusion, glutaraldehyde-stabilized ostrich pericardium tissue shows better mechanical properties than calf tissue. However, calcium accumulation in implanted ostrich tissue is still too high to consider it a much better alternative to bovine pericardium, and anticalcification treatments should be considered.     

Tannic acid-stabilized pericardium tissue: IR spectroscopy, atomic force microscopy, and dielectric spectroscopy investigations

Infrared (IR) spectroscopy, atomic force microscopy (AFM), and dielectric spectroscopy methods were employed to study structural and dynamic changes in the tannic acid (TA)-stabilized pericardium tissue. Chemically stabilized pericardium tissue is widely used in construction of the tissue derived bioprostheses. IR spectra recorded in the range 400-4000 cm-1 allowed us to recognize different types of TA-collagen interactions. Formation of hydrogen bonds between amine as well as amide NH groups from collagen and hydroxyl groups of TA was analyzed. The AFM imaging showed that the stabilization procedure with TA introduces considerable changes in both surface topography and thickness of collagen fibrils as well as in fibril arrangement on the tissue surface. It was found, that these structural changes have an impact on the dielectric behavior of the TA-stabilized tissue. The dielectric spectra for the native and TA-stabilized tissues were measured in the frequency and temperature ranges of 10(-1) -10(7) Hz and 120-270 K, respectively. The dielectric spectra revealed the relaxation process due to orientation of bound water supplemented by the fluctuation of collagen polar side groups. At the temperatures above approximately 210 K, the relaxation due to ion migration process was observed. It was found that both relaxation processes were influenced by the TA-collagen interaction.     

The extension rate independence of the hysteresis in glutaraldehyde-fixed bovine pericardium

Glutaraldehyde-fixed bovine pericardium demonstrates both relaxation of stress and hysteresis during uniaxial loading and unloading. These phenomena suggest viscoelastic behaviour of the material. This study uses analysis of variance to test statistically the hypothesis that the chemically modified pericardium possesses a 'fading memory' for the history of previous load procedures and that the hysteresis is extension rate independent.     

Influence of the selection of the suture material on the mechanical behavior of a biomaterial to be employed in the construction of implants. Part 2: Porcine pericardium

Using a hydraulic stress simulator, the mechanical behavior of the porcine pericardium used in the construction of cardiac valve leaflets was characterized following the same procedure employed with calf pericardium in Part 1 of this study. One hundred fifty pairs of tissue samples were subjected to tensile testing to rupture. One of the two samples from each of 120 pairs (four series of 30 pairs each) was saturated with commercially available threads made of nylon, silk, Prolene or Gore-Tex, while the other sample in each of these pairs was left unsewn. The remaining 30 pairs were employed as controls in which neither of the two samples was subjected to suturing. The sutured tissue samples showed a significant decrease in tensile strength at rupture (range: 11.61 to 21.22 MPa) when compared with unsutured samples (range: 50.80 to 89.45 MPa; p < 0.01). When these results were compared with their equivalent in calf pericardium, no significant differences were observed (the mean values at rupture in calf pericardium ranged between 211.61 MPa and 26.04 MPa). Again, the application of morphological and mechanical selection criteria to ensure the homogeneity of the samples provided excellent fit with respect to the stress/strain curves. The interaction of the different suture materials with the pericardial tissue was also assessed by comparing the mechanical behavior of the sutured samples with that of the control samples. At the working stress of a cardiac valve leaflet, 0.250 MPa, samples sewn with Gore-Tex were found to show the least difference in behavior with respect to the controls, indicating that this material presented the lowest degree of interaction with the pericardium. In conclusion, the suture clearly has deleterious effects on the resistance of both calf and porcine pericardium, which showed no statistically significant differences in terms of resistance to rupture when their respective sutured or unsutured samples were compared, except in the case of porcine pericardium sewn with silk, which presented lower resistance to rupture in all the zones studied. These findings suggest that the hypothesis that porcine pericardium is less resistant is erroneous. The Gore-Tex suture also presented a lower degree of interaction with the porcine pericardium, with values similar to the working stress of a cardiac valve leaflet. This methodology and the results should be evaluated in dynamic studies, such as fatigue testing, that not only confirm the resistance of the material but establish the durability of the samples being assayed.     

不同交联改性牛心包材料的细胞毒性研究

本研究采用培养的人胚肺纤维细胞分析比较戊二醛、环氧１铬交联处理后４％甲醛保存的牛心包材料的细胞毒性。在浸泡前与浸泡１０、２０、３０天的周期中，不同交联处理的牛心包材料与浸出液的细胞增殖抑制指数显示：９１）三种交联剂处理的心包材料均有明显的细胞毒性。（２）随浸泡时期的延长，心包材料与浸出液的细胞毒性逐渐降低。（３）铬交联处理的牛心包材料与浸出液的细胞毒性显著高于其他交联材料。     

Uniaxial and biaxial tensile strength of calf pericardium used in the construction of bioprostheses: biomaterial selection criteria

Using morphological and mechanical criteria and applying a method involving paired samples that is widely employed in epidemiology, we obtained an excellent prediction of the mechanical behavior of the calf pericardium used in the construction of cardiac bioprostheses. The method of selection employed in this study may be a highly useful tool for guaranteeing the mechanical resistance of calf pericardium, with a very low level of error.     

The effects of collagen fiber orientation on the flexural properties of pericardial heterograft biomaterials

Improving cardiac valve bioprostheses (BHV) utilizing heterograft biomaterials requires a better understanding of their mechanical behavior. Flexure is a major mode of deformation for BHV leaflets during valve operation, inducing more complex deformation patterns within the tissue compared to tensile loads. In this study, we investigated the relation between collagen fiber preferred direction and the resulting flexural properties of native and glutaraldehyde-treated bovine pericardium. 20 mm x 4 mm strips were cut from the presorted sheets of bovine pericardium and divided into four groups: two directions of collagen fiber orientation in two groups of native and chemically treated specimens. Specimens were flexed in two different directions using a three-point bending technique (ASAIO J. 45(1999)59) and their flexural mechanical response compared. Results indicated that: (1) the relationship between the applied flexing moment and change of curvature of specimens was non-linear in both native and chemically fixed groups, (2) there were no directional differences in flexural properties when the bovine pericardium is flexed towards either the epi-pericardial or visceral surfaces in both native and chemically fixed specimens, (3) native and chemically fixed bovine pericardium were stiffer when flexed perpendicular to local preferred collagen fiber direction, and (4) chemical fixation increased the flexural rigidity of bovine pericardium. Results of this study indicate that the flexural properties of bovine pericardium are dominated by inter-fiber cross-links as opposed to the stiffness of the collagen fibers themselves. These findings can be used to guide the development of novel chemical treatment methods that seek to optimize biomechanical properties of heterograft biomaterials.     

The telescoping suture--Part 1: Does this technique improve the mechanical behavior of a biomaterial?: Calf pericardium

The authors study the mechanical behavior of calf pericardium employed in the construction of cardiac valve leaflets when subjected to telescoping suture, followed by tensile stress until rupture. One hundred twenty pericardial tissue samples were employed, 60 cut from root-to-apex and another 60 cut in transverse direction. Each of these two groups consisted of 12 control samples that were left unsutured and four sets of 12 samples each that were rejoined by telescoping suture using silk, Prolene, nylon or Gore-Tex., and subjected to tensile stress. At the rupture of the sutured tissues, the tensile stress of the suture materials ranged between 57.54 MPa for the series sewn lengthwise with Gore-tex and 114.08 MPa for the series sewn crosswise with silk. At these levels of stress, the deformation of the suture thread was much less marked than that of the calf pericardium, and internal stresses were produced that were difficult for the biomaterial to absorb. There was a loss of real load in all the sutured series when the observed resistance to rupture, expressed in kilograms, was compared with the estimated value. This loss of resistance did not invalidate the telescoping suture technique since the resistance to rupture was still much greater than that associated with suturing the two edges of the cut pericardium together. This report confirms the deleterious role of the shear force generated in the pericardium by the suture.     

Factor analysis in the evaluation of the relationship between bacterial adherence to biomaterials and changes in free energy.

The relative surface charge and free energy of forty-one coagulase-negative staphylococci were found to be normally distributed; therefore, they can be considered a homogeneous group under strict statistical criteria. The adherence of these bacteria to eight different biomaterials (seven synthetic and one biologic) was found to be independent of charge and variations in free energy during adhesion. Adherence can be explained as a thermodynamic process (free energy decreased with adherence), except in the case of bovine pericardium in which free energy increases. With these biomaterials, a correlation was found between adherence and bacterial charge. Bacterial adherence and bacterial charge correlate with the surface parameters of the biomaterials. This correlation does not occur when the relationships between parameters are evaluated by means of factors analysis, thus indicating the importance of the statistical method selected for the evaluation of bacterial adherence.     

Comparative studies on the relaxing action of several adenosine 5'-triphosphate-sensitive K+ channel openers in pig urethra.

The relaxing effects of the adenosine 5'-triphosphate (ATP)-sensitive K+ channel openers (K(ATP) openers; diazoxide, minoxidil, pinacidil, (+/-)-cromakalim, (+)-cromakalim and (-)-cromakalim) were investigated on the resting tone of pig proximal urethra. In addition, patch clamp techniques were utilized for recording cromakalim-induced ionic currents in cells dispersed from the same urethral region. The (-)-cromakalim-induced relaxation of urethral muscle strips was stable, reversible and reproducible. The rank order of potency regarding of K(ATP) openers in lowering the resting urethral tone was (-)-cromakalim>pinacidil>diazoxide>minoxidil. K(ATP) opener-induced urethral relaxation was suppressed by subsequent application of glibenclamide (1 microM). (+)-Cromakalim (< or =10 microM) did not relax the urethra nor antagonize the (-)-cromakalim-induced urethral relaxation. However, at higher concentrations, (+)-cromakalim (> or =30 microM) caused a small but significant urethral relaxation. In accordance with these observations, the relaxation induced by 5 microM (-)-cromakalim was identical to that induced by 10 microM (+/-)-cromakalim, as expected from a theoretical half potency for (+/-)-cromakalim. In whole-cell recording, (-)-cromakalim and (+) cromakalim (100 microM) activated a glibenclamide-sensitive outward current which was due to the activation of the glibenclamide-sensitive 43 pS K+ channel (K(GS)-43 pS). The potency of (+)-cromakalim to activate K(GS)-43 pS was much weaker than that of (-)-cromakalim. These results indicate that the ability of K(ATP) openers to relax pig urethral smooth muscle can be accounted for by activation of glibenclamide-sensitive K+ channels.     

Gelatinases in soft tissue biomaterials. Analysis of different crosslinking agents

Chemical modification of pericardium-based cardiac valves tends to reduce the relatively high degree of biodegradation and calcification of the implanted bioprostheses. We analysed the tissue properties of pericardium from young calves and pigs after crosslinking with different agents (glutaraldehyde. diphenylphosphorylazide (DDPA), 1-ethyl-3,3-dimethyl-aminopropyl-carbodiimide (EDAC)) and when exposed to anticalcification treatments (chloroform/methanol or ethanol) prior to glutaraldehyde (GA) crosslinking. Protein extraction after tissue homogenisation in the presence of detergents showed that crosslinking using GA or DPPA was much more effective. The amounts of protein extracted from these two groups of chemically modified pericardium were significantly lower: the other modified tissues presented only a slight reduction when compared with untreated tissue. Matrix metalloproteinases- (MMP) 2 and 9 were detected in native pericardium from calf and pig by zymography. While the MMP-9/MMP-2 activity ratio was close to 1 in pig pericardium, it was 8.5-fold higher in bovine tissue. Crosslinking with GA and with DPPA almost completely abolished gelatinase activities, even when equal amounts of solubilised protein were loaded onto the zymograms. Anticalcification treatments followed by GA crosslinking or treatment with EDAC were not as effective in reducing gelatinase activities; but, interestingly, a relative reduction of MMP-9 versus MMP-2 was detected. The presence of these gelatinase activities in pericardium may contribute to the in vivo degradability of pericardium-based cardiac valves.     

Long-term evaluation of five biomaterials for angioplastic enlargement of the pulmonary artery in a young dog model

In 45 young dogs an enlargement angioplasty of the left pulmonary artery was performed using patches made from one of three autologous materials (jugular vein, unmodified pericardium, and glycerolized pericardium) or from two heterologous materials (lyophilized human dura mater and modified bovine carotid artery). Catheterization and angiographic studies performed 5 to 6 months after the operation showed that all patched vessels had remained patent, except in three dogs which had received heterologous implants. The animals were killed 5-24 months after operation (mean weight increase: 84%), and the implants were studied by optical microscopy and morphometry, scanning and transmission electron microscopy, and indirect immunofluorescence with antidog Factor VIII rabbit antiserum. The two heterologous tissues exhibited limited biocompatibility, as estimated from 10 criteria obtained at histologic studies. Conversely, all three autologous biomaterials were characterized by infiltration of noninflammatory cells, near-complete endothelialization, and neosynthesis of structural proteins; infectious foci were very rare or absent. These results suggest that autologous tissues, although deendothelialized at the time of implantation, constitute the most suitable material for patch angioplasty, as far as endothelial triggering, cellularity and resistance to infection are concerned.     

Ostrich pericardium, a biomaterial for the construction of valve leaflets for cardiac bioprostheses: mechanical behaviour, selection and interaction with suture materials.

The mechanical behavior of ostrich pericardium was studied for the purpose of assessing its utility in the construction of bioprosthetic cardiac valve leaflets. The tissue was tested biaxially using a hydraulic simulator that subjected it to increasing stress until rupture. One hundred eighty trials were performed, 36 with unsutured pericardium and four series of 36 trials each with pericardium sutured with silk, Prolene, nylon or Gore-Tex. The samples were tested in pairs from three different pericardial regions. One sample from each pair (the predictive specimen) was assessed according to morphological and mechanical criteria, while the other (the predicted or selectable specimen) was subjected only to morphological analysis. The findings show that ostrich pericardium treated with glutaraldehyde according to standard methods has an excellent resistance to rupture in biaxial testing, withstanding stresses of up to 100 MPa, and never lower than 30 MPa. Its resistance to rupture is lowered by suturing, a loss that is less pronounced when silk sutures are used. The results with Gore-Tex are very homogeneous and the elastic behavior of the pericardium/suture unit appears to be similar to that of unsutured tissue, suggesting that the interaction between the two biomaterials is minor. Similar results were observed in the series sutured with Prolene and nylon. The use of paired samples makes it possible to closely estimate the mechanical behavior of the tissue in a given zone by determining that of its mate. The statistical study shows that this estimation is not conditioned by the suture employed, thus validating this approach and providing more precise criteria for tissue selection.     

Inhibition of the endothelium-dependent relaxation by 18beta-glycyrrhetinic acid in the guinea-pig aorta.

The effect of 18beta-glycyrrhetinic acid (GA), an agent which interferes with gap junction conductivity, on endothelium-dependent relaxation produced by substance P was investigated in isolated aortic rings of the guinea-pig. In nor-adrenaline (NA)-contracted aortic rings, substance P (10(-7) M) induced an endothelium-dependent, transient relaxation. The relaxation was only slightly reduced by the co-application of nitroarginine and diclofenac. When GA (2x10(-5) M) was applied first, it slightly reduced substance P-induced relaxation, and a subsequent co-application of nitroarginine and diclofenac strongly reduced the relaxation. In aortic rings contracted with high-K solution ([K(+)](o) = 29.4 mM), substance P-induced relaxation was reduced by the simultaneous application of GA, nitroarginine and diclofenac, but not by GA alone. In endothelium-denuded aortic rings, GA reduced the threshold concentration of NA required to produce contractions and increased the amplitude of NA-induced contractions. GA increased the amplitude of contraction produced by small increases of [K(+)](o) (<30 mM) but reduced those produced by higher concentrations of [K(+)](o) (>54 mM). In NA-contracted aortic rings, Y-26763, a K(+)-channel opener, could relax muscles with reduced amplitude in the presence of GA. It is concluded that in guinea-pig aortic rings, GA inhibits mainly the EDHF-induced components of endothelium-dependent relaxation. GA also modulated contractions produced by NA or high-K solutions. The possible effects of inhibition of gap junctions by GA on endothelium-dependent relaxation were discussed.     

The involvement of K+ channels and the possible pathway of EDHF in the rabbit femoral artery.

Experiments were designed to characterize the cellular mechanisms of action of endothelium-derived vasodilator substances in the rabbit femoral artery. Acetylcholine (ACh, 10(-8)-10(-5) M) induced a concentration-dependent relaxation of isolated endothelium-intact arterial rings precontracted with norepinephrine (NE, 10(-6) M). The ACh-induced response was abolished by the removal of endothelium. NG-nitro-L-arginine (L-NAME, 10(-4) M), an inhibitor of NO synthase, partially inhibited ACh-induced endothelium-dependent relaxation, whereas indomethacin (10(-5) M) showed no effect on ACh-induced relaxation. 25 mM KCl partially inhibited ACh-induced relaxation by shifting the concentration-response curve and abolished the response when combined with L-NAME and NE. In the presence of L-NAME, ACh-induced relaxation was unaffected by glibenclamide (10(-5) M) but significantly reduced by apamin (10(-6) M), and almost completely blocked by tetraethylammonium (TEA, 10(-3) M), iberiotoxin (10(-7) M) and 4-aminopyridine (4-AP, 5 x 10(-3) M). The cytochrome P450 inhibitors, 7-ethoxyresorufin (7-ER, 10(-5) M) and miconazole (10(-5) M) also significantly inhibited ACh-induced relaxation. Ouabain (10(-6) M), an inhibitor of Na+, K(+)-ATPase, or K(+)-free solution, also significantly inhibited ACh-induced relaxation. ACh-induced relaxation was not significantly inhibited by 18-alpha-glycyrrhetinic acid (18 alpha-GA, 10(-4) M). These results of this study indicate that ACh-induced endothelium-dependent relaxation of the rabbit femoral artery occurs via a mechanism that involves activation of Na+, K(+)-ATPase and/or activation of both the voltage-gated K+ channel (Kv) and the large-conductance, Ca(2+)-activated K+ channel (BKCa). The results further suggest that EDHF released by ACh may be a cytochrome P450 product.     

Effects of ryanodine on relaxation in isolated myocardium from different animal species

Relaxation in mammalian ventricular cardiac muscle is sensitive to the prevailing load. This load dependence of relaxation (LD) can be demonstrated only when an efficient sarcoplasmic reticulum (SR) is present. To define further the role of the SR in LD, we studied contraction and relaxation in cat, rat and frog cardiac muscle after exposure to ryanodine. Ryanodine is a selective inhibitor of calcium release from the SR. This view was confirmed in the present study in single cardiac rat myocytes with functioning SR. Ryanodine did not affect LD in multicellular mammalian myocardium even though it had already significantly depressed contractility, suggesting that calcium release from the SR plays no role in establishing LD. Calcium accumulation in the SR as a consequence of the inhibited release can account for the late depression of LD in the presence of ryanodine.Abbreviations and symbols used LD load dependence of relaxation - LD index for load dependence of relaxation - SE standard error - SR sarcoplasmic reticulum     

Isotonic relaxation in cardiac muscle.

The force-velocity-length determinants of isotonic relaxation were studied in 12 cat papillary muscles. Isotonic relaxation velocity (VL) was found to be a function of total load (preload + afterload), with peak VL increasing to a maximum at loads approximately .3 to .4 Po(L') (Po(L') defined as maximum isometric force developed during a twitch at the experimental length) and falling with increasing loads. Initial muscle length (ML) had no effect on the peak VL with constant load. Increasing the initial length at which isotonic relaxation occurred (LL) decreased peak VL but did not alter the unique length-velocity trajectory at constant load. This unique length-velocity trajectory occurred, despite a wide variation in time during the contraction when peak VL was measured. Increasing Ca++ from 2.5 to 7.5 mM increased peak VL (1.73 +/- .16 to 2.32 +/- .20 ML/s) and shifted the entire length-velocity trajectory toward higher velocities of lengthening. The addition of 10 mM caffeine increased peak VL also (1.67 +/- .18 to 2.54 +/- .20 ML/s) and had a similar effect on the length-velocity trajectory during lengthening as Ca++. Both increased Ca++ and caffeine (10 mM) augmented the maximum VL measured on addition of load.