Surgical isolation of the atrial septum from the atria. Identification of an atrial septal pacemaker.

This is a report of the third in a series of experiments carried out to identify and to determine the reliability of atrial pacemakers below the sinoatrial (SA) node. This information could be useful to the surgeon in planning both corrective atrial operations and direct operations for atrial arrhythmias. In this study, done in dogs, the atrial septum containing the atrioventricular (AV) node was completely separated from the remaining atria. Seven dogs survived for 35 to 116 days. Serial electrocardiograms (ECGs) after operation showed that four of the seven dogs developed a regular junctional rhythm with a rate of 85 beats/min, whereas three of seven developed an irregular junctional rhythm with pauses and bigeminy. In a control study, in which the AV node was disconnected from the His bundle, a slow regular junctional rhythm was produced in all three of the dogs. Thus, in the series of three experiments, this being the report of the last one, a hierarchy of pacemakers below the SA node was identified. The first one, a pacemaker in the low right atrium in the region of the coronary sinus, was the most reliable and was associated with a normal ECG. A second, in the atrial septum, did not develop dominance in all dogs. However, the junctional pacemaker in the His bundle always became dominant after the AV node was disconnected from the His bundle and was associated with a regular but slow rhythm.

Cardiac rhythm following exclusion of the sinoatrial node and most of the right atrium from the remainder of the heart.

In this study two surgical interventions on the right atrium were followed by long-term observation of their effect on the cardiac rhythm. In the first, the sinoatrial (SA) node and a small area of surrounding atrium were excluded from the heart by a circumferential incision. This was followed immediately by an unstable junctional rhythm accompanied by periods of pacemaker arrest. After 14 days a rhythm, indistinguishable from sinus rhythm, became permanently established. On atrial mapping the area of earliest epicardial breakthrough was the low right atrium, indicating that the pacemaker was in this area. In the second procedure, most of the right atrium including the SA node was excluded by an incision from the remainder of the heart, but left in continuity were the left atrium, right atrium in the area about the coronary sinus and inferior vena caval ostium and the atrial septum (areas where the low atrial pacemakers are known to occur), and the ventricles. A permanent junctional rhythm associated with periods of pacemaker arrest was produced. In two dogs permanent atrial fibrillation eventually developed. The area of earliest breakthrough found on epicardial mapping was to the right of the right inferior pulmonary vein, close to the atrial septum. One of the differences between the two interventions was the amount of right atrium remaining between the potential low atrial pacemakers and the atrioventricular (AV) node. It is suggested that summation, a factor needed for the atrial excitation wave to penetrate the AV node, was inadequate in the more extensive intervention. This may explain the failure of a potential low atrial pacemaker to become dominant in the second intervention. The clinical implications of these findings are discussed.

The action of carbon dioxide on the collateral pathways of pulmonary ventilation.

The major collateral ventilatory pathways between the segments of the lobes of the dog are lined with smooth muscle and course from bronchiole to alveolus and from bronchiole to bronchiole. In previous studies, we showed that the channels constricted with metacholine and dilated with epinephrine. Carbon dioxide elevations in the inspired air, as noted in this study, greatly increased the collateral flow, whereas 100 per cent oxygen did not affect it. The greatest dilatation occurred when the inspired carbon dioxide increased from 2.5 to 6 per cent, with little if any further change when increased to 15 per cent. When perfusion of the in situ lung was blocked, or when the lung was excised, carbon dioxide still had a marked bronchodilating effect. This indicates its direct action on smooth muscle. The fact that carbon dioxide acts as a dilator of the collateral channels, independent of neural and humoral influences, confirms its important role in ensuring maximal pulmonary ventilation.

A study of methods for surgical interruption of the His bundle.

The relationship of the atrioventricular (AV) node and His bundle to surrounding structures is described from the viewpoint of the surgeon. A series of experiments were carried out for interruption of atrial to ventricular conduction that included electrocauterization of the His bundle, incision just above the tricuspid annulus to attempt to divide the His bundle, and finally excision of the right atrial wall and the insertion of the atrial septum into the right fibrous trigone to remove all connections from the AV node and atrium to the His bundle. The last procedure was the most satisfactory method, as judged by the certainty of accomplishing both His bundle interruption and interruption at its highest possible point.

Tumor necrosis factor promotes motor functional recovery in crushed peripheral nerve.

The potential benefits of tumor necrosis factor pretreatment in promoting motor functional recovery of peripheral nerve following low load crush injury were examined. Using a specially designed crush device, rat sciatic nerve was subjected to a low load crush injury of 2-h duration. Recombinant murine tumor necrosis factor and saline were intraperitoneally injected into the experimental and control animals, respectively, prior to nerve crushing. Subsequent motor function was evaluated at intervals by measurement of the sciatic functional index. There was significantly (P < 0.05 to < 0.01) more rapid recovery in the tumor necrosis factor pretreated group as compared to the controls between day 14 and day 28. The sciatic functional index in the tumor necrosis factor group improved to -69.3 +/- 5.3 at day 14 and to nearly normal at day 21. In contrast, the sciatic functional index in the control group was -95.5 +/- 3.1% at day 14 and did not approach normal until day 42. Histological results paralleled the functional findings. The results suggest that tumor necrosis factor pretreatment has the potential to attenuate neurostructural damage and promote motor functional recovery in rat peripheral nerve.

Recombinant human glial growth factor 2 (rhGGF2) improves functional recovery of crushed peripheral nerve (a double-blind study).

This in vivo double-blind study evaluated the effect of recombinant human glial growth factor 2 (rhGGF2), a Schwann cell mitogen, on the recovery of motor function of rat sciatic nerve following crush injury. Seventy three rats were divided into three groups. Group I (n=5), sham operated; Groups II (n=34) and III (n=34) received a 100 g crush load for 2 h over a 5 mm segment of the sciatic nerve. Group III was treated with 1 mg/kg rhGGF2, via subcutaneous injection one day before nerve crush and daily for the following four days. Group II received an equivalent volume of saline as a control. Motor functional recovery was assessed by calculating the sciatic functional index (SFI) and the recovery rate of tetanic contractile force of the extensor digitorum longus (EDL) muscle. Recovery of nerve function was evident at day 11 after crush in the rhGGF2-treated animals, whereas the nerves in controls were still paralyzed. The rhGGF2-treated animals showed a significant improvement of the SFI between days 11-21 postoperatively when compared to controls. The isometric tetanic contractile force was stronger in the rhGGF2-treated group than in controls, with a significant difference at 40 to 70 Hz stimulus frequencies on day 4. Correlation analysis showed that tetanic contractile force had a linear correlation with the SFI. Histologic assessment indicated that the rhGGF2-treated animals showed less severe degeneration and earlier robust remyelination of axons than controls. The results suggest that treatment with rhGGF2 is effective in promoting nerve regeneration as seen in measurements of functional recovery and qualitative assessment of nerve morphology. The mechanism of GGF's protective effect may be related to its direct action on Schwann cells, stimulating their mitosis as well as inducing neurotrophic factors essential to neuronal maintenance and repair.

An improved method of recording rat tracks for measurement of the sciatic functional index of de Medinaceli.

The use of paper impregnated with Bromphenol blue for recording normal rat footprints is described. Because paper provides a better grip than X-ray film for the paws of the rats, the resulting tracks make manual scoring easier and more accurate. In addition, the cost of this technique is considerably lower.

Unilateral pulmonary emphysema created by ligation of the left pulmonary artery in newborn puppies.

The left pulmonary artery was ligated in 7 puppies 12 to 24 hours after birth. All were followed by periodic chest roentgenograms. At 6 months of age, ventilatory spaces, split-lung function, and static compliances were determined. Bronchograms and arteriograms were obtained, and histopathology was studied. All animals demonstrated a decrease in left lung size with gross and microscopic changes typical of chronic emphysema. There was impressive bronchial artery hypertrophy. Oxygen consumption and static compliances in the left lungs were depressed. These marked alterations in maturation, as well as the degenerative changes, are believed to be related to an impairment in the nutritive circulation of the lung.

The effect of acute denervation on the microcirculation of skeletal muscle: rat cremaster model.

Although tissue is denervated during replantation of a severed part, tissue transfer, or muscle transplantation, there are few studies concerning the effects of acute denervation on muscle microcirculation. We have described a surgical procedure that totally denervates the rat cremaster muscle. Histological examination of the denervated tissue has given convincing evidence of nerve degeneration and skeletal muscle atrophy, accompanied by electrophysiological evidence of total denervation. The diameters of each component of the microcirculation were measured before and after denervation. Arterioles and arteries ranging in size from 10 to 70 microns in diameter were found to increase significantly in size immediately after acute denervation. Larger arteries and veins did not undergo significant diametrical increases. These findings suggest that total acute denervation significantly increases the diameter of small arteries and arterioles, thereby decreasing the resistance in the arterial bed and increasing blood flow. Since this phenomenon is of limited duration (20 min), it would appear to be ineffective in enhancing reperfusion and oxygenation at the time of reattachment of amputated parts or during vascularized tissue transfers, until methods of prolonging it for several hours or more are found.

Combined effect of acute denervation and ischemia on the microcirculation of skeletal muscle.

Using direct in vivo videomicroscopy and a fluorescein dye technique, reperfusion injury after 3 h of ischemia was studied in the acutely denervated cremaster muscle of the rat. Compared with normally innervated controls, ischemia-induced reperfusion injury was more severe in the denervated group and included a delay of blood flow recovery, vortex formation, edema, hemorrhage, and vessel spasm. Vessel size was reduced at the arteriole and small artery level, and there was a decrease of reactive hyperemia. The injury mechanism may be related to a loss of active vasomotion and vascular response to vasoactive substances after denervation. The results suggest that shortening the ischemia time of denervated tissues may reduce ischemia-induced reperfusion injury. Similarly, given the same ischemia time, improved tissue reperfusion may be expected if the nerve supply is maintained.

Influences of inflation rate and duration on vasodilatory effect by intermittent pneumatic compression in distant skeletal muscle.

Previous study has demonstrated that application of intermittent pneumatic compression on legs can cause vasodilation in distant skeletal muscle at the microcirculation level. This study evaluated the influence of inflation rate and peak-pressure duration on the vasodilatory effects of intermittent pneumatic compression. The cremaster muscles of 50 male rats were exposed and divided into five groups of 10 each. A specially designed intermittent pneumatic-compression device was applied in a medial-lateral fashion to both legs of all rats for 60 minutes, with an inflation rate and peak-pressure duration of 0.5 and 5 seconds, respectively, in group A, 5 and 0 seconds in group B, 5 and 5 seconds in group C, 10 and 0 seconds in group D, and 10 and 5 seconds in group E. Diameters of arterial segments were measured in vessels of three size categories (10-20, 21-40, and 41-70 microm) for 120 minutes. The results showed that the greatest increase in diameter was produced by intermittent pneumatic compression with the shortest inflation rate (0.5 seconds). A moderate increase resulted from compression with an inflation rate of 5 seconds, and no effective vasodilation occurred during compression with the longest inflation rate (10 seconds). When the groups with different inflation rates but the same peak-pressure duration were compared, there was a significant difference between any two groups among groups A, C, and E and between groups B and D. When the groups with different peak-pressure durations but the same inflation rate were compared, compression with a peak-pressure duration of 5 seconds caused a generally similar degree of diameter change as did compression without inflation at peak pressure. The findings suggest that inflation rate plays an important role in the modulation of distant microcirculation induced by intermittent pneumatic compression whereas peak-pressure duration does not significantly influence the vasodilatory effects of the compression. This may be due to the fact that rapid inflation produces a significant increase in shear stress on the vascular wall, which stimulates vascular endothelium to release nitric oxide, causing systemic vasodilation.

Protective effect of hypothermia on contractile force in skeletal muscle.

The clinical success of limb replantation and tissue transfer is partly dependent on the duration of ischemia experienced by the amputated part. This study focused primarily on the damage that occurs during this ischemic period. An experimental system was implemented that allowed the observation of contractile function in totally isolated skeletal muscle after ischemia. Contractile function was selected as an indicator of ischemic damage because normal function is the ultimate goal of replantation. All experiments were performed on the rat extensor digitorum longus. The muscles were subjected to ischemic periods of 1.5, 3.0, and 5.0 hours and were stored in either a hypothermic (4 degrees C) or a room-temperature (23 degrees C) environment during the ischemic interval. After the ischemic period, all muscles were transferred to a tissue bath and were subjected to contractility testing, followed by fatigue testing. In both groups, muscle function decreased as the ischemic interval was increased. A significant difference in function between the normal control and the muscles of both ischemic groups implied that ischemic injury had occurred in the hypothermic and room-temperature muscles, even with the relatively short 1.5-hour ischemic interval. After each ischemic interval however, the hypothermic muscles produced significantly greater contractile force than the room-temperature muscles in both the contractility and the fatigue tests. After 1.5 hours of ischemia, the contractile force in the hypothermic group was about three times as great as that observed in the room-temperature group. These results indicated that muscle function after a period of totally isolated ischemia is protected by hypothermic preservation. They also support the advisability of storage of amputated parts and free muscle flaps in hypothermic environments before replantation even after relatively brief intervals of ischemia.

Effects of immobilization on Achilles tendon healing in a rat model.

The aim of this study was to evaluate the effects of immobilization and mobilization on the functional and biomechanical recovery of injured Achilles tendons. Male Sprague-Dawley rats were allocated randomly into four groups: (a) sham operation, (b) division only (surgical transection of the Achilles tendon without immobilization), (c) "dummy" external fixation (division of the Achilles tendon and application of Kirschner wires), and (d) rigid external fixation (division of the Achilles tendon and immobilization with Kirschner wires connected by two triangular frames). All procedures were performed on the right lower limb; the left, uninjured, lower limb served as an internal control. Kirschner wires and external fixators were removed on day 12. Functional performance was determined from measurements of hind pawprints of rats walking preoperatively and on postoperative days 1, 3, 5, 7, 9, 11, 13, and 15. On day 15, the animals were killed and biomechanical evaluations were performed on both the injured and the uninjured Achilles tendon constructs. No functional or mechanical deficits were observed in the sham-operation group. Animals subjected to division of the Achilles tendon had an initial functional deficit that returned to near normal by day 15. The application of Kirschner wires was associated with an impairment of the functional performance of the rat as well as of the mechanical properties of the tendon-bone constructs. Immobilization by connection of the Kirschner wires to an external frame had an additional, highly significant (p < 0.001) detrimental effect on the functional and mechanical recovery of Achilles tendon-calcaneal complexes.

Achilles tendon healing: a correlation between functional and mechanical performance in the rat.

The pathogenesis and treatment of rupture of the Achilles tendon remain a source of controversy. This study presents the results of a biomechanical, functional, and morphological evaluation of a group of rats that had division and repair of the Achilles tendon. A total of 46 rats were used: 18 for biomechanical testing, 18 for functional evaluation, and 10 for histology. Morphological examination revealed an early inflammatory response with loose connective tissue formation that was replaced gradually by fibroblasts and a collagenous matrix. The functional evaluation (Achilles functional index [AFI]) was made from measurements of the hind pawprints of walking rats. Division and repair of the Achilles tendon produced a significant functional impairment (mean [+/- SEM] AFI = -87 +/- 8; p < 0.001), which gradually improved with healing time. The load to failure for the repaired tendons consistently improved with healing time, in a manner similar to the functional recovery. The average deformation (repair/control) varied considerably and was not related to healing time. The stiffness of the repaired tendons increased with healing time and was 60% of the corresponding control side by day 15. The major finding of this study was a strong correlation between the AFI and the failure load of the healing tendon-bone constructs (250-300 g group, r = 0.97, p < 0.001; 325-375 g group, r = 0.96, p < 0.001).

An experimental canine model of osteonecrosis: characterization of the repair process.

Osteonecrosis of the femoral head was induced in dogs by a process of deep freezing, accompanied by stripping of the soft-tissue attachments from the femoral neck and intertrochanteric area, in an effort to develop an experimental model to study treatment modalities for avascular necrosis of the hip. Immediate uniform necrosis was created in a defined area. Thereafter, spontaneous healing originated mainly from the adjacent viable bone by migration of undifferentiated mesenchymal tissue into the necrotic bone, genesis of fibrosis, and, finally, formation of new bone. Osteogenesis occurred primarily through intramembranous ossification without a preexisting template. Quantitative measurements showed a difference in the rates of ingrowth and revascularization of the necrotic area during the fibrotic and osteogenic phases of the healing. The de novo osteogenesis was slower than the migration of fibrosis. These findings indicate that, without osteotomy, we can reproducibly create necrosis of bone in a defined area of the proximal femur and induce a reparative process that incompletely heals the defect. Although it does not fully simulate the human disorder of osteonecrosis, our experimental surgical model provides a basis for further laboratory investigation into the management of avascular necrosis of the femoral head.

Calcitonin gene-related peptide and reperfusion injury.

Calcitonin gene-related peptide is a potent intrinsic vasodilator, can induce prostacyclin release, and may inhibit membrane lipid peroxidation. This study examines the effect of calcitonin gene-related peptide on vessel diameters, capillary perfusion, and contractile function of skeletal muscle after 4 or 5 hours of ischemia and during immediate reperfusion using the rat cremaster muscle model. Forty-two male rats were used; half of these received 0.2 ml of 10(-7) M calcitonin gene-related peptide after 0, 15, and 30 minutes of reperfusion, while the other half received normal saline as a control. By means of intravital videomicroscopy, the diameters of 10 vessels per muscle were measured prior to ischemia and during reperfusion. The fluorescein filling area was determined at 15, 30, and 60 minutes of reperfusion. After 1 hour of reperfusion, muscle function was examined in vitro by quantifying the contractile response to electric field stimulation of the muscles in an organ bath system. There was a significant increase in the diameter of the arterioles, but not the small arteries, at every time point from 10 to 60 minutes of reperfusion. The fluorescein filling area was increased in treated muscles at every time point. Contractile function was not significantly preserved. In light of the ability of calcitonin gene-related peptide to relieve vasospasm and improve capillary perfusion, it may be useful in reducing reperfusion injury in the future.

Gentamicin distribution from a collagen carrier.

Local delivery of antibiotics by a degradable carrier has the potential for high local antibiotic levels and avoids systemic toxicity. Intravenous access, renal function monitoring, and subsequent surgical removal may not be required when degradable local delivery modalities are used. This study examined the in vivo elution of gentamicin from processed bovine collagen (type I) in 66 adult White rabbits. Collagen impregnated with gentamicin (3 mg/kg) was implanted into the vastus lateralis, and data were collected from 15 minutes to 28 days after implantation. Local tissue biopsies were taken a minimum of 2 mm from the implantation site. The gentamicin was released into the local tissue and averaged more than 3,800 micrograms/ml during the initial 4 hours after implantation. Local levels fell to 6.90 +/- 5.22 micrograms/ml at 24 hours and subsequently were 2.70 +/- 1.75 micrograms/ml or more through day 28. Serum levels reached an average peak of 4.04 +/- 1.75 micrograms/ml at 5 hours after implantation, decreased after the initial 24 hours, and subsequently were less than 0.41 +/- 0.20 microgram/ml through day 28. Collagen impregnated with gentamicin proved to be an effective degradable carrier of gentamicin in the healthy rabbit; it provided local tissue concentrations above the minimum inhibitory concentration and serum concentrations below levels associated with systemic toxicity for 28 days after implantation.

Protective effect of 21-aminosteroid pretreatment in peripheral nerve low-load crush injury in mature and immature rats.

The effects of U-74006F (tirilazad mesylate), a 21-aminosteroid antioxidant, on injured peripheral nerve were studied. Twenty-two immature and 44 mature rats were divided equally into two groups. The experimental group received two injections of 3 mg/kg of U-74006F at a 2 hour interval. The control group received the same volumes of a citrate buffer. A 5 mm segment of the sciatic nerve was subjected to a crush load of 100 g for 2 hours. Motor function (sciatic functional index) was assessed to day 48 postoperatively. There was total paralysis of the crushed limb in all rats the first week after crushing. The experimental group had a statistically significant improvement in motor function compared with the controls on days 14, 21, 25, and 28 for the mature rats and on days 11 and 14 for the immature rats. The mature controls attained complete recovery on day 42 and had a significantly slower recovery rate than the immature controls, which had recovered fully by day 25. The recovery rates were almost similar among mature and immature groups pretreated with U-74006F, both of which had fully recovered motor function by day 28. The results indicate that pretreatment with U-74006F can significantly promote peripheral nerve function after low-load crush injury and that the age of the animal influences the rate of peripheral nerve recovery.

Motor functional and morphological findings following end-to-side neurorrhaphy in the rat model.

Nerve repair cannot always be achieved by the conventional end-to-end technique. This study evaluated the functional recovery of nerves repaired with end-to-side neurorrhaphy in a rat model. The right peroneal nerves of 80 female rats were transected and divided into four groups. In group A, the nerve ends were separated and remained unrepaired; in group B, the distal peroneal ends were directly sutured to the epineurium of the tibial nerves in end-to-side fashion; in group C, the distal ends were sutured through an epineurial window at the repair site in end-to-side fashion; and in group D, the nerve ends were reconnected by the traditional end-to-end technique. Evaluation included gait analysis by calculation of a peroneal functional index, measurement of contractile function of the extensor digitorum longus muscle, wet weight of the extensor digitorum longus, and histological examination. The findings of this study suggested the following: (a) end-to-side neurorrhaphy allows effective motor functional recovery, demonstrated by earlier improvement of the peroneal functional index, stronger muscle contractile function, greater muscle weight, and higher density of regenerated axons compared with unrepaired nerves; (b) removal of the epineurium of the donor nerve at the nerve coaptation site increases the effectiveness of end-to-side neurorrhaphy, but the epineurium appears to be a partial barrier to axonal regeneration; (c) removal of the epineurium does not affect the structure and function of the donor nerve; and (d) end-to-end repair achieved the best functional recovery among the four groups; therefore, end-to-side repair should be considered as a potential alternative only when no proximal nerve is available.

Interleukin-1 beta promotes functional recovery of crushed peripheral nerve.

This study was conducted to elucidate the role of the cytokine interleukin-1 beta on peripheral nerve recovery following crush injuries of two different magnitudes. Eighty-eight female rats were divided into four groups. A 5-mm segment of the right sciatic nerve was subjected to a 100-g crush load for 2 hours in the rats in Groups A1 and B1 or to a 15,000-g crush load for 10 minutes in the rats in Groups A2 and B2. The rats in Groups A1 and A2 received 10 microg/100 g body weight human recombinant interleukin-1 beta intraperitoneally 48, 24, and 1 hours before the nerve injury. The rats in Groups B1 and B2 were treated with an equal volume of normal saline solution with identical schedule guidelines. Walking-track tests (sciatic functional index) performed at intervals until 56 days after the crush and measurements of the contractile force of the extensor digitorum longus muscle made until 28 days were used to evaluate functional recovery of the nerve. During the second week after injury, the rats treated with interleukin-1 beta (A1) had an earlier recovery on the walking track than did those treated with saline solution (B1); this difference reached significance (p < 0.05) at day 11. Although Group A2 demonstrated a trend toward earlier recovery compared with Group B2, there was no significant difference between the two groups. After low or high-load crush injury, tetanic contractile forces were greater in the rats treated with human recombinant interleukin-1 beta than in those treated with saline solution. The results suggest that treatment with human recombinant interleukin-1 beta before crush injury can promote function in the peripheral nerve after the injury. However, the mechanisms that underlie the observed beneficial effects are not completely understood and only speculations can be made.