Diss. (sammanfattning) Uppsala : Univ., 2001. / Härtill 5 uppsatser.

Thesis (Ph. D.)--University of Wisconsin--Madison, 1950. / Typescript. Vita. With this are bound: The determination of glutamic acid dehydrogenase in tissue homogenates / by J.H. Copenhaver, Jr., W.H. McShan, and Roland K. Meyer. Reprinted from Journal of biological chemistry, vol. 183, no. 1 (Mar. 1950), p. 73-79 -- Succinic dehydrogenase and anaerobic glycolysis in livers of diabetic rats / Elva G. Shipley, Roland K. Meyer, J.H. Copenhaver, Jr. and W.H. McShan. Reprinted from Endocrinology, vol. 46, no. 3 (Mar. 1950), p. 334-337. Includes bibliographical references (leaves 40-44).

Kwan Hon Pong. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2000. / Includes bibliographical references (leaves 115-156). / Abstracts in English and Chinese. / Abstract --- p.i / 摘要 --- p.iii / Abbreviations --- p.v / Acknowledgment --- p.vii / Chapter 1. --- Introduction / Chapter 1.1 --- Diabetes mellitus - general introduction --- p.1 / Chapter 1.1.1 --- Animal models of diabetes --- p.5 / Chapter 1.1.2 --- Streptozotocin-induced diabetes (SID) --- p.6 / Chapter 1.1.2.1 --- Mechanism of the diabetogenic effect of STZ --- p.7 / Chapter 1.1.2.2 --- Administration of STZ --- p.9 / Chapter 1.2 --- Impairment of cognitive function in diabetes mellitus --- p.9 / Chapter 1.3 --- Common mechanisms suggested in diabetic neuropathy --- p.15 / Chapter 1.3.1 --- Polyol pathway activation --- p.15 / Chapter 1.3.2 --- Redox potential alterations --- p.16 / Chapter 1.3.3 --- Nonenzymatic glycation --- p.17 / Chapter 1.3.4 --- PKC alteration --- p.18 / Chapter 1.4 --- Do the common mechanisms of neuropathy induced the cognitive impairment in diabetes --- p.18 / Chapter 1.5 --- Structure and function of the hippocampus --- p.20 / Chapter 1.6 --- The definition and mechanism of learning and memory --- p.21 / Chapter 1.7 --- The mechanisms underlying the early and late phases of LTP in hippocampus --- p.23 / Chapter 1.7.1 --- Perforant and schaffer collaterals pathways --- p.23 / Chapter 1.7.2 --- Mossy fibre pathway --- p.24 / Chapter 1.7.3 --- Late phase of LTP in hippocampus --- p.25 / Chapter 1.8 --- GABAergic interaction in hippocampal plasticity --- p.25 / Chapter 1.9 --- The objective of the project --- p.27 / Chapter 1.10 --- Hypothesis --- p.27 / Chapter 1.10.1 --- The initial role of glutamate receptors --- p.28 / Chapter 1.10.2 --- Involvement of putative retrograde messengers --- p.30 / Chapter 1.10.3 --- The role of GABA receptors --- p.37 / Chapter 1.10.4 --- The role of the CREB --- p.40 / Chapter 2. --- Materials and methods / Chapter 2.1 --- Animals --- p.43 / Chapter 2.1.1 --- Induction of diabetes mellitus --- p.43 / Chapter 2.1.2 --- Insulin therapy --- p.45 / Chapter 2.1.3 --- Sample collection --- p.46 / Chapter 2.2 --- Isolation of total RNA --- p.47 / Chapter 2.3 --- Quantitation of total RNA --- p.51 / Chapter 2.4 --- Reverse transcription --- p.53 / Chapter 2.5 --- PCR --- p.54 / Chapter 2.5.1 --- Preparation of PCR --- p.54 / Chapter 2.5.2 --- Purification of PCR product --- p.60 / Chapter 2.5.3 --- Confirmation of PCR products by DNA sequencing --- p.61 / Chapter 2.5.4 --- PCR analysis --- p.62 / Chapter 2.5.4.1 --- Quantitation of cDNA --- p.62 / Chapter 2.5.4.2 --- Radioactive PCR --- p.65 / Chapter 2.5.4.3 --- cDNA gel electrophoresis --- p.66 / Chapter 3. --- Results / Chapter 3.1 --- Ionotropic glutamate receptor subtypes --- p.72 / Chapter 3.1.1 --- Non-NMDA receptors --- p.72 / Chapter 3.1.1.1 --- AMPA receptors --- p.72 / Chapter 3.1.1.2 --- Kainate receptors --- p.72 / Chapter 3.1.2 --- NMDA receptors --- p.76 / Chapter 3.2 --- Metabotropic glutamate receptor subtypes --- p.79 / Chapter 3.2.1 --- Group I subtype --- p.79 / Chapter 3.2.2 --- Group II subtypes --- p.79 / Chapter 3.3 --- Synthases of retrograde messengers --- p.79 / Chapter 3.4 --- Calcium-related receptors --- p.82 / Chapter 3.5 --- "GABA receptor subtypes (Aαl-4,BRla)" --- p.85 / Chapter 3.6 --- Glutamic acid decarboxylase (GAD) --- p.88 / Chapter 3.7 --- Enzyme genes related to CREB dephosphorylation --- p.88 / Chapter 3.8 --- Effect of insulin therapy on ionotropic glutamate receptor subtypes --- p.91 / Chapter 3.9 --- Effect of insulin therapy on metabotropic glutamate receptor subtypes --- p.91 / Chapter 3.10 --- Effect of insulin therapy on synthases of retrograde messenger --- p.91 / Chapter 3.11 --- Effect of insulin therapy on GAB A receptor subtype --- p.91 / Chapter 4. --- Discussion / Chapter 4.1 --- SID on Glutamate receptor subtypes --- p.96 / Chapter 4.2 --- SID on Calcium-related receptors --- p.105 / Chapter 4.3 --- SID on Synthases of retrograde messengers --- p.106 / Chapter 4.4 --- SID on GABA receptor subtypes --- p.109 / Chapter 4.5 --- SID on enzyme genes related to dephosphorylation of CREB --- p.111 / Chapter 4.6 --- Effect on insulin therapy on gene expression in hippocampus --- p.113 / Chapter 5. --- References --- p.115

Made available in DSpace on 2015-07-22T20:50:33Z (GMT). No. of bitstreams: 0 Previous issue date: 2006-12-31 / O objetivo do presente estudo foi avaliar a participação do sistema renina angiotensina através de avaliações “ in vivo” e “in vitro” na disfunção cardíaca induzida pelo diabetes experimental por estreptozotocina. Foram utilizados ratos Wistar machos (250-300g) divididos em 4 grupos: Controle Saudável (n=9), Controle Enalapril, (n=7, 1m/Kg), Diabético (n=7, STZ, 50mg/Kg ev), e Diabético + Enalapril (n=7). Nos estudos “in vivo” foi avaliada a função ventricular de forma não invasiva pelo ecocardiograma e invasiva pela cateterização do ventrículo esquerdo (VE) no período basal e após sobrecarga de volume e a pressão arterial (PA) de forma direta nos animais acordados. As dosagens bioquímicas incluíram medida da enzima de conversão da angiotensina (ECA) no soro e coração dos ratos, e em cultura primária de fibroblastos cardíacos tratados com glicose no meio de cultura (25 mM). O diabetes induziu perda de peso corporal gradativa ao longo das 4 semanas e hiperglicemia, alterações que não foram atenuadas pelo tratamento com enalapril. Observou-se prejuízo em parâmetros morfométricos (aumento da cavidade do VE, diminuição da espessura do septo intraventricular e da parede posterior do VE) e de função contrátil (redução da fração de ejeção e da velocidade de encurtamento circunferencial; aumento da velocidade média do picoE, do tempo de desaceleração da onda E e do tempo de relaxamento isovolumétrico) no coração dos animais diabéticos (15 e 30 dias). O tratamento com enalapril atenuou tais prejuízos. Na análise funcional invasiva do VE em 30 dias, no período basal, o grupo diabético apresentou redução da pressão sistólica do VE (controle: 134 ± 13 vs diabético 113 ± 14 mmHg*) e da contratilidade do VE, avaliada pela +dP/dt (controle: 9229 ± 1225 vs diabético: 6565 ± 1610 mmHg/seg *) e pela - dP/dt (controle: - 6845± 1002 vs diabético: - 4745 ± 1557 mmHg/seg*), além de aumento da pressão diastólica final do VE (PDF) (controle: 4,98 ± 0,98 vs diabético: 7,36 ± 0,5 mmHg*) quando comparados ao grupo controle. O tratamento com enalapril não modificou esses parâmetros de função do VE no período basal. Além disto, as diferenças observadas entre o grupo controle e o grupo diabético foram mantidas após um protocolo de sobrecarga de volume. Entretanto, os animais diabéticos tratados com enalapril após a sobrecarga de volume apresentaram medidas de função diástólica do VE (PDF e –dP/dt) semelhantes aos valores do período basal, diferentemente do grupo diabético, no qual, ao final do protocolo de sobrecarga, observou-se valores dobrados de PDF em relação ao período basal. A análise direta dos sinais de PA (Windaq, 2KHz) nos animais acordados evidenciou hipotensão e bradicardia nos grupos diabéticos, tratados ou não com enalapril, em relação aos grupos controles. No soro dos animais diabéticos a atividade da ECA utilizando o substrato ZPhe-HHL aumentou em torno de 50% em relação aos animais controles, no entanto, o tratamento com enalapril não inibiu essa atividade. O oposto foi observado no tecido cardíaco: no grupo diabético a atividade da ECA reduziu em torno de 25%, enquanto, o controle tratado não teve inibição da ECA quando comparados ao controle. O grupo diabético tratado com enalapril obteve uma inibição da ECA em torno de 50%(substrato ZPhe-HHL) em relação aos controles tratados ou não. O mesmo perfil foi observado com o substrato HHL no soro e coração. Entretanto, observou-se uma ativação maior da ECA pelo substrato ZPhe, tanto no soro como no coração em relação ao substrato His Leu. Na análise da expressão proteica da ECA por “western blotting” no coração observou-se uma expressão protéica da ECA aumentada nos animais diabéticos e diabéticos tratados em relação aos controles com os dois anticorpos utilizados: alto peso molecular (136KDa) e de baixo peso (69KDa). Os resultados de expressão protéica aumentada da ECA, foram acompanhados de redução da atividade da enzima no tecido cardíaco. A atividade bioquímica da ECA analisada em cultura de fibroblastos cardíacos estava aumentada em torno de 85% após tratamento com glicose. Concluí-se que o diabetes experimental por estreptozotocina induziu prejuízo em parâmetros cardíacos morfométricos e de função sistólica e diastólica de forma semelhante ao que vem sendo documentado em humanos. Porém a intervenção terapêutica com enalapril tanto em 15 quanto em 30 dias de diabetes atenuou essas disfunções sem, no entanto, promover normlaização da PA e frequência cardíaca. O fato da prejuízo da função do VE observada nos animais diabéticos ter sido inibida pelo tratamento com enalapril sugere que a ativação do sistema renina angiotensina tem um papel importante nas disfunções cardiovasculares do diabetes. / The aim of the present study was to elucidate the role of renin angiotensin system, by “in vivo” and “in vitro” evaluation, on cardiac dysfunction induced by streptozotocin experimental diabetes. Male Wistar rats (250-300g) were divided in 4 groups: control (n=9), control + Enalapril, (n=7, 1mg/Kg), diabetic (n=7, STZ, 50mg/Kg ev), and diabetic + Enalapril (n=7). In vivo studies included: echocardiography as a non invasive tool for ventricular function evaluation and catheterization of left ventricle (LV) to evaluate invasively. The last one was performed in basal condition and after a volume overload protocol. Arterial pressure (AP) was measured directly in awake animals. Biochemistry dosage included angiotensin converting enzyme (ACE) activity in serum, heart and primary cardiac fibroblast culture treated with glucose (25 mM). Diabetes induced hyperglicemia and progressive body weight loss during the protocol. These alterations were not attenuated by enalapril treatment. There were impairment on morphometric (increased LV cavity, reduced intraventricular septum thickness and LV posterior wall thickness) and contractile function parameters (reduced ejection fraction and velocity of circumferential shortening; increased mean E peak velocity, E wave desacceleration time and isovolumetric relaxation time) in the diabetic animalshearts (15 and 30 days). Enalapril treatment attenuated these impairments. In the invasive LF evaluation (30 days), in the basal period, diabetic group presented diminished LV systolic pressure (control: 134 ± 13 vs diabetic 113 ± 14 mmHg*) and LV contractility, measured by +dP/dt (control: 9229 ± 1225 vs diabetic: 6565 ± 1610 mmHg/seg*) and by -dP/dt (control: - 6845± 1002 vs diabetic: - 4745 ± 1557 mmHg/seg*), and also enhanced LV end diastolic pressure (EDP) (control: 4,98 ± 0,98 vs diabetic: 7,36 ± 0,5 mmHg*) as compared to control group. Enalapril treatment did not modify these LV functional parameters in the basal period. Furthermore, the differences observed between control and diabetic groups were maintained after the volume overload protocol. However, enalapril treated diabetic animals presented LV diastolic function parameters (PDF e –dP/dt) after the volume overload similar to their resting values. Differently, the diabetic group showed twice PDF values after volume overload in comparison to it basal PDF values. Direct AP signals measurements (Windaq, 2KHz) in awake animals evidenced hypotension and bradycardia in diabetic groups, treated or not with enalapril, when compared to control groups. Diabetic animals’ serum ACE activity using ZPhe-HHL substrate was 50% increased when compared to control animals; however, enalapril treatment did not inhibit this activity. The opposite was observed in heart tissue: ACE activity reduced 25% in diabetic group and control treated group did not present ACE inhibition in relation to control group. Enalapril treated diabetic group showed ~50% ACE (substrate ZPhe-HHL) inhibition in comparison to treated or untreated control groups. Similar profile was evidenced with ACE substrate HHL in serum and heart. However, it was observed higher ACE substrate ZPhe activation, in both, serum and heart, in comparison to ACE substrate His Leu. Heart ACE protein expression by “western blotting” was increased in diabeticgroups (treated and untreated) in relation to control group with two antibodies: high molecular weight (136KDa) and low molecular weight (69KDa). The increased ACE protein expression was accompanied by heart reduced ACE activity. Glucose treatment increased ~85% ACE activity in primary cardiac fibroblast cultures. In conclusion, streptozotocin experimental diabetes induced impairment in morphometric cardiac parameters and in systolic and diastolic function. Similar condition has been reported in humans. Despite of enalapril therapeutic intervation, in 15 and 30 days, had attenuted these dysfunctions in diabetics, it did not induce AP or heart rate normalization. The inhibition of LV function impairments by enalapril treatment suggests that renin angiotensin system activation plays an important role in the diabetic cardiovascular dysfunctions . / TEDE / BV UNIFESP: Teses e dissertações

A prominent finding in the diabetic rat heart is a decrease in the rate at which the ventricular muscle can contract and relax. Since cardiac sarcoplasmic reticulum is thought to be intimately involved in muscle contraction and relaxation we studied the ability of diabetic rat cardiac sarcoplasmic reticulum to transport Ca²⁺ . Hearts were obtained from female Wistar rats 7, 30, 42, and 120 days after the induction of diabetes by a single i.v. injection of either alloxan (65 mg/kg) or streptozotocin (60 mg/kg). At all Ca²⁺ concentrations tested (0.2 μM-5.0 μM free Ca²⁺) cardiac sarcoplasmic reticulum obtained from 42 and 120 day diabetic rats showed a significant decrease in the rate of ATP-dependent tns-oxalate facilitated ²⁺ransport. This was accompanied by a decrease in Ca²⁺ -ATPase activity. The levels of long chain acylcarnitines associated with the microsomal sarcoplasmic reticulum preparation from 42 and 120 day diabetic rats were significantly higher than those present in sarcoplasmic reticulum from control rats. Palmitylcarnitine, the most abundant of the long chain acylcarnitines, in concentrations < 7 μM was found to be a potent time-dependent inhibitor of both Ca²⁺ transport and Ca²⁺ -ATPase in both control and diabetic rat sarcoplasmic reticulum preparations; inhibition of Ca²⁺ transport was found to be more marked in the control preparations. This would indicate that a degree of inhibition produced by the high endogenous levels of palmitylcarnitine may already be present in the diabetic rat heart preparations. Cardiac sarcoplasmic reticulum prepared from acutely diabetic rats (7 days) did not show any decrease in Ca²⁺ transport ability. Levels of long chain acylcarnitines associated with the microsomal preparation enriched in sarcoplasmic reticulum were also unchanged. Insulin treatment of diabetic rats could significantly increase the ability of cardiac sarcoplasmic reticulum to transport Ca²⁺, although at the time period obtested (30 days) the SR Ca²⁺ transport activity was only slightly depressed as compared to control. Insulin treatment also resulted in a slight, but non-significant, lowering of the levels of long chain acylcarnitines associated with the sarcoplasmic reticulum microsomal preparations. These findings suggest that the alteration in sarcoplasmic reticulum function in chronically diabetic rats may be due to the buildup of cellular long chain acylcarnitines which inhibit sarcoplasmic reticulum Ca²⁺ transport. The absence of any significant change in Ca²⁺ transport activity or levels of long chain acylcarnitines at 7 and 30 days suggests that the alterations in 42 and 120 day diabetic rats must be of gradual onset. Cardiac sarcoplasmic reticulum is known to be regulated by a number of factors, among them calmodulin, cAMP-dependent protein kinase, and K⁺. Since Ca²⁺ transport activity in cardiac sarcoplasmic reticulum from chronically diabetic rats is depressed, the role that these regulators play was investigated. Calmodulin (0.61 μM), cAMP (10 μM) plus cAMP-dependent protein kinase (0.2 mg/0.5 ml), and K⁺ (0-110 mM) all stimulated Ca transport in both control and streptozotocin-treated diabetic rats to the same degree. This suggests that the depression observed in sarcoplasmic reticulum function from diabetic rats is not due to altered regulation by these putative mediators of Ca²⁺ uptake. A number of studies suggest that carnitine administration may lower myocardial levels of long chain acylcarnitines in the diabetic rat. Therefore, D,L-carnitine (1 g/kg/day, orally) was administered to 120 day diabetic rats for a 30 day period. The elevated levels of long chain acylcarnitines normally seen in diabetic rats were significantly reduced in the diabetic rats administered carnitine. Carnitine administration, however, could not reverse the previously noted depression in diabetic rat heart function, as measured on an isolated working heart apparatus. In an effort to prevent the onset of the diabetic cardiomyopathy D,L-carnitine was administered (3 g/kg/day, orally) 3 days after the induction of diabetes for a 42 day period. As previously mentioned, sarcoplasmic reticulum Ca²⁺ transport activity was depressed in diabetic rats, as compared to control rats, at all free Ca²⁺ concentrations tested (0.1 μM-3.5 μM). Similarly, sarcoplasmic reticulum levels of long chain acylcarnitines were significantly elevated in these diabetic rats. The diabetic rats treated with carnitine did not show any depression in Ca²⁺ transport activity; long chain acylcarnitine levels were also similar to control. The carnitine-treated diabetic rats, however, showed no improvement in heart function compared to untreated-diabetic rats. These data suggest that although the long chain acylcarnitines are inhibiting cardiac sarcoplasmic reticulum function in chronically diabetic rats other factors must also be contributing to the depression in heart function. / Pharmaceutical Sciences, Faculty of / Graduate

The nature and extent of myocardial ischemic injury (Mil) produced either by coronary artery ligation/reperfusion or by injection of isoproterenol (ISO) was studied in the 10-week alloxan-diabetic rabbit. Prior to the induction of ischemic injury, investigation of the left ventricles of the diabetic rabbit after 10-weeks revealed significant magnesium depletion and inhibition of myofibrillar and sarcoplasmic reticulum ATPase activities. In addition, the activity of the lysosomal enzyme, N-acetyl-β-glucosaminidase was significantly increased in diabetic left ventricular homogenates. Ultrastructural studies revealed significant lipid and glycogen accumulation, dilatation of the sarcoplasmic reticulum and damage to the mitochondria in left ventricles of the diabetic animals. Administration of ISO to both control and diabetic animals resulted in atrial tachycardias and ventricular fibrillation. The severity of the arrhythmias and the overall mortality was the same in both groups of animals. Serum analyses revealed significantly greater increases in blood glucose, free fatty acids, total cholesterol and creatine kinase activity in the ISO-treated diabetic animals relative to ISO-treated controls. ISO treatment of both control and diabetic animals produced similar increases in heart weight, left ventricular weight and myocardial water content. Analyses of various subcellular organelle marker enzyme activities indicated a significantly greater decrease in the K⁺ ,Ca²⁺ -stimulated sarcoplasmic reticulum ATPase of ISO-treated diabetic animal hearts. In addition, significantly greater increases in Ca and hydroxyproline and decreases in the levels of ATP were evident in the ISO-treated diabetic animal hearts. Ultra-structural studies revealed significant damage to the mitochondria in both ISO-treated control and diabetic hearts, the magnitude of the damage being greater in the diabetic animals. Mitochondria from both groups of animals showed swelling and fragmentation, myofibrils appeared as a homogeneous mass and did not show the characteristic Z-lines. Glycogen depletion and lipid accumulation was observed in both groups of animals. In addition, both groups of animals showed amorphous dense bodies in the mitochondria after ISO-treatment. After 40-minutes occlusion of the left circumflex coronary artery followed by 60-minutes of reperfusion, hemodynamic measurements revealed significant decreases in the left ventricular and systemic arterial pressures in the diabetic animals relative to controls. Analyses of subcellular organelle enzymes from the ischemic tissue revealed that sarcolemmal Na⁺ ,K⁺ -ATPase, mitochondrial ATPase and sarcoplasmic reticulum ATPase activities were decreased after coronary occlusion in both control and diabetic animals. However, upon reperfusion, unlike the control, no recovery of the mitochondrial ATPase was observed in the diabetic animals. In addition, a further depression of both the sarcolemmal and sarcoplasmic reticulum ATPase activities were seen in the diabetic animals compared to controls on reperfusion. Ion measurements revealed a significant accumulation of calcium in both control and diabetic animals, the magnitude of the increase being greater in the diabetic animals. Similarly, both tissue ATP levels and the ability of the mitochondria to generate ATP were depressed in the diabetic animals as compared to controls following coronary artery occlusion and reperfusion. Following coronary artery ligation and reperfusion, the diabetic animals showed a significantly higher incidence of ventricular fibrillation and cardiogenic shock as compared to controls. Ultrastructural studies revealed myocardial damage to both control and diabetic hearts following coronary artery ligation and reperfusion. However, the diabetic myocardium showed a higher incidence and frequency of hypercontraction bands, an increase in the amorphous dense bodies and slightly greater damage to the mitochondria. Coronary artery ligation in conscious control, 6 and 12 week-diabetic rats resulted in post-ligation arrhythmias (especially ventricular fibrillation), the incidence of which was much greater in the diabetic animals. The mortality rate of 12-week diabetic rats undergoing coronary ligation was 100% within 1-7 minutes following ligation. No differences in occluded or infarcted zones of the surviving 6-week diabetic and control rats were detected. Analyses of ionic composition revealed a significant magnesium deficiency in the diabetic hearts as compared to controls. These data indicate that the diabetic animals show a greater susceptibility of the myocardium to ischemic injury. Although numerous metabolic and chemical alterations are present in the diabetic myocardium, it is possible that magnesium deficiency may be a factor determining the higher incidence of arrhythmias and ischemic injury in diabetic animals. / Pharmaceutical Sciences, Faculty of / Graduate

Diabetes is known to result in a large number of alterations which affect various systems and organs. One of the more prominent disorders associated with diabetes is that of cardiac disease. Clinically, diabetics suffer from morbidity and mortality of cardiac origin to a greater extent than the nondiabetic population. Various functional studies have also revealed that the efficiency of diabetic hearts to function as pumps is lower than that of normal hearts. Experimentally, myocardial function of either rats or dogs made diabetic with either streptozotocin (STZ) or alloxan has been studied and a depression clearly demonstrated in both the species. The abnormalities of cardiac function in experimental diabetes are accompanied by depression of various enzyme systems in the heart. These include the ability of the sarcoplasmic reticulum (SR) to take up calcium; the myosin and actomyosin ATPase activities; and the Na⁺, K⁺ ATPase activity. All these changes can be prevented and reversed by insulin treatment suggesting that the myocardial problems seen in STZ or alloxan diabetic animals are due to diabetes and not direct toxicities of the drugs. It is not known whether the beneficial effects of in vivo insulin treatment are due to its direct myocardial effects or whether they are secondary to its effects mediated via normalisation of metabolism in diabetic animals. Thus, in the first part of the present investigation, we examined the direct effects of insulin on hearts from either control or diabetic rats using the isolated working heart preparation. Rats made diabetic with STZ (55 mg/kg) were sacrificed either 3 days or 6 weeks after induction of the disease and their hearts isolated and perfused in the working heart mode. Glucose concentrations varying from 5mM to 20mM were used in the perfusion medium, either in the presence or absence of insulin (5mU/mL). Left ventricular function was expressed as left ventricular developed pressure (LVDP) and the rates of contraction and relaxation (positive and negative dP/dt respectively) at various left atrial filling pressures. Three days after injecting STZ into rats, the animals exhibited hypoinsulinemia, hyperglycemia and their body weights although not significantly different from those of control animals, tended to be lower than the body weights of controls. Animals treated in this manner did not exhibit depression of cardiac function when compared with the myocardial function of control rats. Hearts from control rats exposed to regular insulin in the presence of 5mM glucose exhibited values of contractility which were significantly greater as compared with those obtained from control rat hearts not exposed to the hormone. When insulin was perfused along with a higher concentration of glucose (10mM), function of control rat hearts was affected to a significant extent. As opposed to the effects on control rat hearts, insulin failed to increase contractility in hearts from 3 day diabetic rats when either 5 or 10mM glucose was used in the perfusion medium. The study was then repeated using animals which had been diabetic for six weeks. At the time of sacrifice, these animals were hypoinsulinemic, hyperglycemic and weighed significantly less than their age-matched controls. Analysis of cardiac function revealed a significant depression in diabetic rats as compared with controls. Increasing glucose concentrations from 5 to 20mM in the perfusion medium did not affect the function of either control or diabetic rat hearts. Perfusion with regular insulin increased contractility in control rat hearts; the increase in contractility was not affected by increasing the glucose concentration from 5 to 10mM. However, contractility of diabetic rat hearts was not affected by insulin perfusion when either 5 or 10mM glucose was used in the perfusion medium. In order to eliminate the possibility of involvement of glucagon (which may contaminate commercial insulin preparations) in the effects of insulin on control rat hearts, part of the study was repeated using glucagon - free insulin. While the glucagon - free insulin increased contractility in control rat hearts, diabetic rat hearts were not affected. These results are identical to those obtained with regular insulin, suggesting that the effects of insulin observed were due to insulin itself. Although insulin treatment prevents and reverses diabetes - induced myocardial alterations in the rats, due to its widespread metabolic effects, it is not a good tool for investigating the specific factors which cause the cardiac abnormalities. In addition, a major problem with insulin treatment clinically is the fact that hypoglycemia can be associated with it, inadequate control occurs in some diabetics and secondary complications, such as myocardial problems, occur despite insulin treatment. It is thus desirable to have treatments which selectively affect certain aspects of diabetes so that the suspected underlying causes can be corrected specifically and their significance in causing the myocardial problems assessed. It would also be useful to have drug treatments which could either substitute for insulin or could be used in addition to the peptide. We have thus studied the effectiveness of certain treatments in preventing diabetes - induced myocardial alterations. The first one used was methyl palmoxirate, a fatty acid analog which is reported to reduce blood glucose levels in diabetic rats and dogs. The glucose - lowering effect is mediated via inhibition of fatty acid metabolism due to inhibition of carnitine acyl transferase resulting in inhibition of acyl carnitine formation and eventually inhibition of fatty acid transport across the mitochondrial membrane. Rats were treated with the drug (25mg/kg/day p.o.) three days after they were injected with either STZ or buffer. The treatment was carried out for 6 weeks and cardiac performance was then assesed. Untreated and treated diabetic rats were hypoinsulinemic, hyperglycemic and hyperlipedemic at the time of sacrifice. Cardiac function, which was depressed in diabetic animals, was still depressed despite the methyl palmoxirate treatment. However, the ability of the myocardial sarcoplasmic reticulum (SR) to take up calcium, which was depressed in diabetic rats, was normal in treated diabetic rats. Also, the levels of long chain acyl carnitines (LCAC) in the myocardial SR were normalised by methyl palmoxirate treatment in diabetic rats. In an effort to normalise diabetes - induced myocardial alterations in rats, we then attempted a combination of either methyl palmoxirate or carnitine (as both can prevent the depression of SR calcium uptake) with thyroid hormone treatment (as it can normalise myosin ATPase depression in diabetic rat hearts). The treatment protocol was identical to that described above (30µg/kg/day s.c. T₃ was used). Although the general features of both control and diabetic animals were not affected by either of the combination treatments, cardiac dysfunction in diabetic rats was prevented by methyl palmoxirate and T₃ treatment. Carnitine and T₃ treatment, on the other hand, affected the function of diabetic rat hearts only at the lower left atrial filling pressures. These results suggest that the combination treatment of methyl palmoxirate and T₃ affect parameters besides SR calcium uptake and myosin ATPase. This is because the combination of carnitine and T₃, which also supposedly affects same parameters as the other combination, could not prevent the myocardial alterations. One of the possible reasons for the effectiveness of the combination of methyl palmoxirate and T₃ could be that animals treated with methyl palmoxirate derived at least part of their metabolic energy (especially at higher left atrial filling pressures) from glucose and thus reduced the oxygen demand at higher filling pressures as opposed to the untreated diabetic rat hearts which depended completely on fatty acids for their metabolic energy demands. / Pharmaceutical Sciences, Faculty of / Graduate

This document only includes an excerpt of the corresponding thesis or dissertation. To request a digital scan of the full text, please contact the Ruth Lilly Medical Library's Interlibrary Loan Department (rlmlill@iu.edu).

This document only includes an excerpt of the corresponding thesis or dissertation. To request a digital scan of the full text, please contact the Ruth Lilly Medical Library's Interlibrary Loan Department (rlmlill@iu.edu).

This document only includes an excerpt of the corresponding thesis or dissertation. To request a digital scan of the full text, please contact the Ruth Lilly Medical Library's Interlibrary Loan Department (rlmlill@iu.edu).