Effects of methylphenidate (Ritalin) on mammalian myocardial ultrastructure.

Previous observations have indicated lamellated ultrastructural lesions in the myocardium of a patient treated with methylphenidate (Ritalin) hydrochloride (MPH). A causal relationship between MPH exposure and these membranous changes was tested in the myocardium of rats and mice. Following injection of varying doses of MPH for different periods, myocardial ultrastructure was examined and lesions were quantified by stereological techniques. Myocardial tissue also was stained using techniques selective for acid phosphatase and for sarcoplasmic reticulum to identify possible pathogenetic mechanisms. MPH induced membrane accumulations and lamellations which were not membrane-bound and did not react for acid phosphatase, but stained positively for sarcoplasmic reticulum. Both lesions were highly focal, surrounded by normal appearing myocardial tissue. Lamellations were evident at the earliest timepoints examined and appeared to occur without lysosomal involvement. Lesions were still apparent 12 weeks after terminating MPH. These data suggest that MPH may have persistent, cumulative effects on the myocardium.

Cocaine-induced myocardial ultrastructural alterations and cardiac output responses in rats.

Cocaine use has been associated with profound functional and pathological myocardial responses in otherwise asymptomatic humans, yet a number of individuals appear to tolerate large doses of the drug. This study was designed to determine whether there is a relationship between the differential effects of cocaine administration on cardiovascular responses and on the development of cardiomyopathies in rats. After instrumentation for determination of cardiac output, conscious, freely moving rats were treated with cocaine (5 mg/kg) or saline intravenously twice daily for 14 days before removing the myocardium for analysis. Although most cardiovascular responses were similar, cocaine administration elicited consistent decreases in cardiac output in some rats, whereas others showed little change or an increase. While little change was evident at low magnification, electron microscopy revealed diffusely distributed myocardial lesions including focally dilated sarcoplasmic reticulum and myofibrillar derangement, early signs of mitochondrial alterations, and foci of myocardial fibrosis. The incidence of these alterations was greater in rats with a decrease in cardiac output. We also observed these lesions in a subset of rats treated with cocaine without cardiac output instrumentation. These data represent the first evidence that there is a relationship between cocaine-induced functional and pathological alterations and that rats, like humans, may be differentially sensitive to these effects.

Effects of dilation with a balloon catheter on the endothelium of the internal thoracic artery.

Controlled dilation of the internal thoracic artery with a balloon catheter has been reported to effectively treat intraoperative arterial spasm. It has been shown in laboratory animals that dilation of the internal thoracic artery at prescribed shear force levels will not cause intimal damage. Using scanning electron and light microscopy, we have examined the effects of calibrated balloon dilation on the endothelium of the human internal thoracic artery. In 10 patients with bilateral internal thoracic artery grafting, the artery was dilated with a Fogarty IMAG balloon catheter (Baxter Healthcare Corporation, Edwards Division, Santa Ana, Calif.) that was withdrawn at tensions of 20 or 30 gm. Arterial segments and nondilated control specimens were prepared for scanning electron microscopy. The intimal surface of each internal thoracic artery was evaluated by assigning a score (from 0 to 3) to 10 examined scanning electron microscopy fields; subsequently the arterial tissue was viewed by light microscopy with paraffin-embedded sections stained for elastic tissue. Arteries were obtained from three additional patients so that the microscopic appearance of the arteries could be observed after rough manipulation or removal of the balloon without shearing. The results of this study are as follows: (1) By scanning electron microscopy, dilated internal thoracic arteries yielded consistently higher scores than the control arteries, reflecting severe, tension-dependent alterations of the endothelium, which included marked desquamation of endothelial cells, with extensive areas of complete denudation and pronounced attachment of platelets to these areas; (2) endothelial injury occurred by inflation alone, without shearing by the inflated balloon; (3) by light microscopy, the internal thoracic arteries showed (a) fenestrations of the internal elastic lamina with occasional transmigration of smooth muscle cells through these gaps and (b) foci of intimal thickening without overt atherosclerotic lesions. We conclude that the endothelium of human internal thoracic arteries is highly vulnerable to balloon dilation, which can severely injure the intimal surface. For this reason we prefer not to include this procedure in our protocol for preparing the internal thoracic artery.

Use of the inferior epigastric artery as a free graft for myocardial revascularization.

From March 1990 through January 1991, 47 patients undergoing myocardial revascularization had one (37) or both (10) inferior epigastric arteries (IEA) used as a conduit for bypass with 62 distal anastomoses. The internal thoracic artery (ITA) was used bilaterally in 41 patients and unilaterally in 6 with 100 distal anastomoses. Five patients had a single saphenous vein graft. In total, 167 anastomoses (3.55 per patient) were performed. Single IEA grafts were harvested through a paramedian incision and bilateral grafts, a midline incision. Harvest time was 36.5 minutes for IEA grafts and 29.6 minutes for ITA grafts (p less than 0.0001). Graft length was 11.9 cm for IEA grafts and 16.5 cm for ITA grafts (p less than 0.0001). Distal graft diameter was 2.0 mm for IEA grafts and 2.1 mm for ITA grafts (p less than 0.01). Graft flow was 49.7 mL/min for IEA grafts and 48.7 mL/min for ITA grafts. Microscopic assessment of segments of both the IEA and ITA from 14 patients revealed similar internal elastic laminae and an equal number of fenestrations. Combined intimal and medial thickness was comparable in both conduits. Medial elastic tissue was more prominent in ITA grafts and lacking in eight of the 14 IEA grafts. Gross plaque formation was noted in the proximal 1 to 3 cm of 50% of IEA grafts, but the lumen was not compromised and microscopic thickening was minimal. An unexpected finding was medial calcifications (Mönckeberg's disease) in two of the 14 IEAs without associated atherosclerosis. There was one hospital death, one abdominal wound infection, and one instance of fat necrosis superficial to the sternum.(ABSTRACT TRUNCATED AT 250 WORDS)

Mural repair following obliteration of aneurysms. Part III: Pathomorphological effects of Nd:YAG laser on aneurysm obliteration.

Twenty-four experimental aneurysms were created in rat carotid arteries using the venous pouch technique. Four to twelve weeks later, these aneurysms were totally obliterated by the external application of Nd:YAG laser. The aneurysmal dome and neck were both exposed to the laser using low amplitude (1-2 W) at continuous or repeat intervals (0.3 sec on/0.1 sec off). Small aneurysms were easily obliterated without external signs of necrosis, whereas aneurysms larger than 2 mm required complete coagulation resulting in a charcoal-like appearance. At varying intervals (30 min, 7 days, 3 weeks, 6 weeks), the obliterated aneurysms were harvested and evaluated using the scanning electron microscope and standard histological techniques. The results indicate that the effects of the laser on the aneurysm and parent vessel are similar to those encountered following the application of bipolar coagulation with massive coagulation necrosis of the aneurysmal neck and dome. Notably, however, the extension of this process onto the parent vessel involving especially the endothelium surrounding the aneurysm orifice (commonly seen with bipolar coagulation) is minimal following laser coagulation. There appears to be a protective effect on the parent vessel endothelium by blood flow through the vessel. On this basis, it appears that the laser may be an alternative method of aneurysmal coagulation; the use of laser entails less manipulation of the aneurysm. The Nd:YAG laser may be a useful adjunct in the surgical obliteration of clinical cerebral artery aneurysms, especially small ones.

Altered angioarchitecture in selected areas of brains with Alzheimer's disease.

It was the aim of this study to determine, qualitatively and quantitatively, alterations in the blood vessels of brains removed postmortem from patients with Alzheimer's disease (AD), and to compare these findings with the appearance of cerebral blood vessels in a group of individuals without brain disorders. Celloidin sections of brain tissue from four cerebral areas, pre-frontal (Brodmann's area 9), basal forebrain, sensorimotor, and hippocampus, were subjected to an alkaline phosphatase reaction to facilitate the evaluation of the vascular distribution. The vascular density in five sections was determined by counting the number of vascular intersections with a microscopic test grid of 100 squares; ten fields per section were examined in this manner. Analysis of 16 AD and 6 control brains, showed that there was a striking and statistically significant reduction in the vascular net density specifically in the basal forebrain region and the hippocampus of AD brains. In addition, vessels in the AD brains exhibited extensive topographical changes, such as kinking and looping. These results indicate that modifications in vascular density are present in AD brains with a marked regional specificity.

In search of the "perfect" anastomosis.

Forty-five end-to-side microvascular anastomoses were completed in rat carotid arteries of 0.7-0.8 mm diameter (anastomosing the distal end of the left common carotid to the side of the right common carotid). For comparison both 10-0 and 11-0 sutures were utilized in different anastomotic techniques: interrupted, direct-continuous, and diagonal-continuous sutures, plus total mural thickness vs. partial mural thickness (piercing only the adventitia and outer media, excluding the intima). Anastomoses were evaluated for patency and scanning electron microscopic appearance after 10 to 12 weeks. The results indicate complete patency in all anastomoses. Ultrastructural observations revealed nearly normal intimal appearance in the partial medial technique and only minimal evidence of intimal injury in the other techniques. It is concluded that 100% patency can be obtained regardless of suture size or anastomotic technique. The most important factor in anastomotic patency is the operator's technical skill.

Modification of anionic sites in myocardial capillary basal laminae of diabetic rats.

Capillary basal laminar thickening is a distinctive feature of diabetic microangiopathy; however, the mechanism responsible for this abnormality remains to be clarified. Recent reports have described a reduction in the distribution of anionic sites in diabetic glomerular basement membranes, with the suggestion that this reduction may generate a compensatory synthesis of basal laminar constituents, causing laminar thickening. In order to provide additional information, the character and distribution of the basal laminar anionic profile were examined in the myocardium of diabetic rats. Diabetes mellitus was induced in 14 rats by injection with streptozotocin, ip; 6 rats served as controls. Myocardial tissue was subjected to Charonis' procedure for the demonstration of anionic sites with the cationic electron-dense dye, ruthenium red, following the sacrifice of the animals at intervals up to 11 months after the induction of the diabetes. The tissues were then processed routinely for electron microscopic examination. A total of 20 electron micrographs, at magnifications of 13,000x and 33,000x, were obtained from each rat for the quantitation of anionic sites. A length measuring 6 micron along each basal lamina was utilized for determining the number of anionic loci. Results of this study show that (1) the number and size of anionic sites in myocardial basal laminae is reduced in diabetic rats, (2) this decrease becomes more pronounced with prolongation of the diabetes, (3) it is detectable prior to the demonstration of basal laminar thickening by electron microscopy, and (4) enzyme digestion treatments indicated that heparan sulfate proteoglycan is the essential stainable component of the anionic sites. These findings provide evidence that the laminar anionic profile is altered in the diabetic myocardium and support the view that this abnormality constitutes a significant initial event in the pathogenesis of basal laminar thickening.

Mural repair following obliteration of aneurysms: II. Pathomorphology of treated aneurysms.

Mural vascular repair following experimental and clinical aneurysm obliteration was studied using the scanning electron microscope and standard histological techniques. Two of the more frequently utilized clinical obliteration techniques, clipping and coagulation, were examined. A chronological description of the mural reparative process reveals that clipping is superior to coagulation in most instances, that coagulation results in satisfactory obliteration of the aneurysmal orifice only in very selected circumstances, and that a combination of coagulation and clipping may be the best overall obliteration technique.

Collagenous abnormalities in the heart of the tight-skin mouse.

The tight-skin (TSK) mouse, a possible animal model for scleroderma, has multiple abnormalities including increased dermal thickness, cardiomegaly, emphysematous lungs, and an enlarged skeleton. Previous investigations have demonstrated an increased collagen and glycosaminoglycan (GAG) content in the skin and lungs of these mice. The present correlative investigation of the biochemical and ultrastructural properties of the heart in the TSK mouse also revealed an increased presence of collagen. Analysis of collagen types in the TSK heart showed there was a shift in the ratio of type I: type III: type V from the normal values. Over 90% of the collagen was type I, while both types III and V were decreased in this organ. The ultrastructural examination of the left ventricle demonstrated extensive accumulations of perivascular and intercellular edema fluid, foci of myocytolysis, and areas of moderately increased collagen deposits within interstitial sites. These findings suggest that an increased collagen deposition (type 1) may be a contributing factor to cardiac enlargement in the TSK mouse.

The effects of ionophores on steroidogenesis and morphology of avian granulosa cells.

Granulosa cells, isolated by collagenase digestion from the mature ovarian follicle of laying hens, were incubated in the presence of two ionophores, lasalocid (X537A) and ionomycin, to determine their effects on basal and stimulated steroidogenesis, as well as their effects on various cell parameters including DNA, RNA, and protein synthesis. Both ionophores caused a dose-dependent inhibition of agonist-promoted progesterone production and, in the presence of calcium, a small but significant increase in basal output of progesterone. Whereas the conversion of pregnenolone to progesterone was unaffected by the ionophores, the activity of cholesterol side-chain cleavage enzyme was inhibited in a dose-related manner. Both ionophores decreased cellular levels of ATP and inhibited the incorporation of radioactively-labeled precursors into DNA, RNA, and proteins. Morphologically, ionophore-treated cells showed swelling of the rough endoplasmic reticulum. Similar morphological changes were also observed in cells treated with oligomycin, a known metabolic inhibitor. These results suggest that the ionophores lasalocid and ionomycin impair release of energy and thereby exert the principal cause of the inhibited steroidogenic response by granulosa cells to a variety of agonists.

Mural repair following obliteration of aneurysms: production of experimental aneurysms.

One hundred thirty-two experimental rat carotid artery aneurysms were produced using a variety of methods, in order to determine which of these approaches should be employed in the development of a satisfactory laboratory model. The gross appearance and pathomorphology of aneurysms created by vein pouch and partial arteriotomy methods most closely resembled those features reported in clinical intracranial aneurysms.

Thyrotoxic myopathy in mice: accentuation by a creatine transport inhibitor.

To demonstrate the importance of creatine and phosphocreatine in skeletal muscle during periods of metabolic stress, thyrotoxicosis was induced in mice fed the creatine transport inhibitor, beta-guanidinopropionic acid (beta-GPA). Adding 2% of beta-GPA to the diet of normal mice inhibited weight gain and caused a 75% reduction of creatine and phosphocreatine concentrations in skeletal muscle. Addition of 0.25% or 2% of thyroid powder to the diet of normal mice was associated with hyperactivity, cardiomegaly, and a high mortality rate. Superimposing thyrotoxicosis on mice already depleted of creatine and phosphocreatine resulted in degeneration of muscle fibers. These results indicate that high concentrations of creatine and phosphocreatine are essential for the maintenance of muscle integrity during periods of metabolic stress.

Mitigation of an anthracycline-induced cardiomyopathy by pretreatment with razoxane: a quantitative morphological assessment.

A quantitative evaluation of structural modifications was undertaken in the myocardium of daunorubicin (DNR)-treated and razoxane (RZ)-protected mice. BDF1 mice were injected with DNR, 15 mg/kg; a second group of mice was subjected to the same conditions but, in addition, received a pretreatment of RZ, 200 mg/kg. Representative cubes of myocardial tissue were processed for viewing with the electron microscope. Five hundred myocardial cells in each group were examined for the presence of lesions which had been categorized as early, moderate, or advanced. Contrasting the total number of demonstrable lesions in each group revealed a statistically significant reduction of 38% in abnormalities present in RZ-protected mice. By category, RZ-pretreated mice showed a mitigation in the appearance of early and moderate alterations and a striking reduction in the incidence of advanced, irreversible lesions. These results indicate that the cardiomyopathy associated with DNR administration can be ameliorated by pretreatment with RZ; this protective effect is markedly exerted by preventing the development of severe, irreversible lesions in the murine myocardium; the initial, non-transient structural alteration subsequent to DNR-exposure appears to affect the myocardial sarcoplasmic reticulum.