O agonista beta-adrenérgico ractopamina (RAC) modifica a composição da carcaça suína por aumentar a massa muscular e reduzir a deposição de gordura. O objetivo neste estudo foi avaliar os efeitos do tempo de fornecimento de RAC sobre o desempenho, concentração de ureia plasmática (CUP), características de carcaça e expressão gênica dos receptores beta- adrenérgicos (beta-AR) e das isoformas da cadeia pesada de miosina (MyHC) em suínos em terminação. Oitenta suínos, machos castrados (PV inicial = 69,42 ± 1,24 kg), foram utilizados em um experimento em blocos completos casualizados com cinco tratamentos, oito repetições por tratamento e dois animais por unidade experimental (baia). Os tratamentos consistiram de rações sem RAC (controle) ou com 10 ppm de RAC fornecidas por 7, 14, 21 ou 28 dias préabate. O PV individual e o consumo de ração por baia foram obtidos para determinar o ganho diário de peso (GDP), o consumo diário de ração (CDR) e a conversão alimentar (CA). Amostras de sangue foram coletadas para determinação da CUP. No final do experimento, os animais (PV final = 102,46 ± 1,44 kg) foram abatidos e amostras de pelos e do músculo Longissimus dorsi coletadas. As carcaças foram avaliadas 24 horas post-mortem. As amostras de pelos foram utilizadas para detecção da mutação no gene do receptor de rianodina do tipo 1 (RYR1). A expressão gênica dos beta-AR (subtipos beta1 e beta2) e das isoformas MyHC (I, IIa, IIx/d e IIb) foi quantificada nas amostras de músculo. As análises estatísticas foram realizadas apenas com os animais homozigotos dominantes para a mutação no gene do RYR1. O aumento no período de fornecimento de RAC não afetou (P > 0,05) o PV final, o GDP e o CDR, porém resultou em melhora linear (P < 0,01) na CA. Melhoras (P < 0,05) nas médias semanais de GDP e CA foram observadas durante os primeiros 21 dias de fornecimento de RAC, no entanto, o crescimento animal declinou (P < 0,05) na 4ª semana de tratamento. A CUP apresentou efeito quadrático (P < 0,01) com o aumento na duração do fornecimento de RAC. Houve aumento linear (P <= 0,01) no peso da carcaça quente, na profundidade do músculo Longissimus dorsi, na área de olho de lombo e na relação carne:gordura com o aumento na duração do tratamento com RAC. Não foram detectados efeitos da RAC (P > 0,05) sobre a expressão gênica dos beta1-AR e das isoformas MyHC IIa e MyHC IIx/d, porém o aumento no período de fornecimento de RAC tendeu a reduzir linearmente (P = 0,08) a expressão gênica dos beta2-AR. Embora os níveis de RNAm da isoforma MyHC I tenham sido reduzidos linearmente (P < 0,01), a expressão gênica da isoforma MyHC IIb aumentou linearmente (P < 0,01) com o aumento na duração do tratamento com RAC. Portanto, as melhores respostas de desempenho e carcaça ocorreram quando a RAC foi fornecida por 21 e 28 dias, respectivamente. Além disso, o agonista alterou a expressão gênica das isoformas MyHC, e é possível que a ação da RAC esteja relacionada com a população de beta2-AR. / The beta-adrenergic agonist ractopamine (RAC) modifies the swine carcass composition by increasing muscle mass and decreasing fat deposition. The objective in this study was to evaluate the effects of RAC feeding duration on performance, plasma urea nitrogen (PUN) concentration, carcass traits, and gene expression of beta-adrenergic receptors (beta-AR) and myosin heavy chain (MyHC) isoforms in finishing pigs. Eighty barrows (initial BW = 69.42 ± 1.24 kg) were used in a randomized complete block design experiment with five treatments, eight replicates per treatment, and two animals per experimental unit (pen). The dietary treatments consisted of diets containing no RAC (control) or 10 ppm RAC fed for 7, 14, 21, or 28 days before slaughter. Individual pig BW and pen feed disappearance were obtained to determine average daily gain (ADG), average daily feed intake (ADFI), and feed to gain ratio (F:G). Blood samples were collected for determination of PUN concentrations. At the end of the experiment, pigs (final BW = 102.46 ± 1.44 kg) were slaughtered and hair and Longissimus dorsi muscle samples collected. The carcasses were evaluated 24 hours postmortem. Hair samples were used to detect the mutation of the ryanodine receptor type 1 (RYR1) gene. Gene expression of beta-AR (beta1- and beta2-subtypes) and MyHC isoforms (I, IIa, IIx/d, and IIb) was quantified in the muscle samples. Statistical analyses were performed using only the homozygous dominant pigs for the RYR1 gene mutation. Increasing RAC feeding period did not affect (P > 0.05) final BW, ADG, and ADFI, but resulted in a linear improvement (P < 0.01) in F:G. Average weekly improvements (P < 0.05) in ADG and F:G were observed during the first 21 days of RAC feeding, however, animal growth declined (P < 0.05) in the 4th week of treatment. The PUN concentrations showed a quadratic effect (P < 0.01) as RAC feeding duration increased. There were linear increases (P <= 0.01) in hot carcass weight, Longissimus dorsi muscle depth, loin eye area, and muscle to fat ratio as RAC treatment duration increased. No effects of RAC feeding (P > 0.05) were detected for beta1-AR and for isoforms of MyHC IIa and MyHC IIx/d gene expression, but increasing RAC feeding period tended to linearly decrease (P = 0.08) beta2-AR gene expression. Even though mRNA levels of MyHC I isoform decreased linearly (P < 0.01), gene expression of MyHC IIb isoform increased linearly (P < 0.01) as RAC treatment duration increased. Therefore, greater growth and carcass responses occurred when RAC was fed for 21 and 28 days, respectively. Furthermore, the agonist altered the MyHC gene expression and the RAC action may be related to the beta2-AR population.

Os hormônios tireoidianos (HT) executam um papel lipolítico importante no Tecido Adiposo Branco (TAB), sendo este efeito mediado por meio do aumento da expressão de receptores beta adrenérgicos na membrana do adipócito, o que aumenta a sensibilidade deste tecido as catecolaminas. Sabe-se que os principais efetores da ação lipolítica nesse tecido são a lipase hormônio sensível (LHS) e a lipase dos triglicerídeos dos adipócitos (ATGL), as quais hidrolisam os triglicerídeos em ácidos graxos e glicerol. Além disso, outros componentes estão envolvidos na atividade lipolítica, como as perilipinas, proteínas estas que envolvem a gota de gordura, formando uma barreira contra a ação da LHS e ATGL, de modo que precisam ser fosforiladas para que a LHS e ATGL possam exercer seu efeito lipolítico. Considerando: (a) a importância do tecido adiposo na homeostase energética e como fonte de citocinas, as quais estão relacionadas com a sensibilidade tecidual à insulina; (b) que a função e o metabolismo do tecido adiposo variam com a sua distribuição regional, e (c) que as ações lipolíticas dos HT, importantes reguladores da homeostase energética, têm sido muito pouco exploradas, pretendemos investigar, em ratos, (i) se os HT interferem na expressão da LHS, ATGL, perilipina A e dos receptores beta3 adrenérgicos no tecido adiposo branco, e (ii) se essas ações diferem nos distintos depósitos de gordura, o que poderia ampliar o campo de conhecimento sobre os efeitos lipolíticos destes hormônios e a nossa compreensão sobre a contribuição deles nas complicações associadas à obesidade e suas co-morbidades. / Thyroid hormones (TH) play an important lipolytic role in white adipose tissue (WAT). This effect is mediated by increased expression of beta-adrenergic receptors on adipocytes membrane, which increases the sensitivity of that tissue to catecholamines. It is known that the main effectors of the lipolytic action in WAT enzymatic activity, especially: hormone sensitive lipase (HSL) and adipose triglyceride lipase (ATGL), which hydrolyze triglycerides into fatty acids and glycerol. In addition, other components are involved in the lipolytic activity, such as perilipin. These proteins support the fat droplet, forming a protective barrier against HSL and ATGL action. Considering: (a) the importance of adipose tissue in energy homeostasis and as a source of cytokines which are related to insulin tissue sensitivity; (b) function and metabolism of adipose tissue vary with their regional distribution; and (c) lipolytic actions of HT, important regulators of energy homeostasis, have been little explored, we investigated in rats with hypothyroidism and submitted to T3 treatment: (i) TH effects on the expression of hormone sensitive lipase (HSL), adipose triglyceride lipase (ATGL),perilipin A and beta-3 adrenergic receptors in WAT, and (ii) if this action are different on subcutaneous and visceral fat depot. This study has increased our understanding about the contribution of these hormones on WAT metabolism and metabolic disease as obesity.

O objetivo desta pesquisa foi avaliar os efeitos do período de administração de cloridrato de zilpaterol (CZ) e do tempo de confinamento no desempenho, características de carcaça, rendimento de cortes cárneos comerciais e na qualidade de carne de bovinos Nelore. Foram utilizados 96 bovinos machos não castrados da raça Nelore, com peso vivo inicial de 377 ± 25,1 kg. O delineamento utilizado foi o de blocos completos casualizados em esquema fatorial 4 × 2, sendo quatro períodos de administração com CZ (0, 20, 30 ou 40 dias) e dois tempos de confinamento (90 ou 117 dias). O período de suplementação com CZ (8,33 mg/kg de matéria seca total) iniciou-se 23, 33 e 43 dias antes do abate. Nos últimos três dias que antecederam o abate todos os animais foram alimentados com a dieta sem CZ, respeitando o período de carência do produto. O contrafilé (músculo Longissimus thoracis et lumborum) de todas as meias-carcaças esquerdas foi amostrado para as análises de qualidade de carne. Com exceção da área do olho de lombo obtida por ultrassom (P = 0,02) e área do olho de lombo medida na carcaça resfriada (P = 0,05), não houve interação entre período de administração de CZ e tempo de confinamento (P > 0,10). O aumento do período de suplementação com CZ proporcionou aumento linear da eficiência alimentar (P < 0,01), peso de carcaça quente (P < 0,01), rendimento de carcaça quente (P < 0,01) e rendimento de carne desossada (P < 0,01). Os bifes de contrafilé de animais suplementados com CZ apresentaram maior conteúdo percentual de proteína (P < 0,01), enquanto que a espessura de gordura subcutânea (P = 0,04) e o conteúdo percentual de lipídios intramusculares (P < 0,01) diminuíram com administração de CZ. O valor percentual de ácidos graxos poliinsaturados totais aumentou (P = 0,01) com a administração de CZ, sendo atribuído principalmente ao aumento percentual do ácido linoleico (18:2 cis-9, cis-12; P = 0,01) e ácido araquidônico (20:4 n-6; P < 0,01). A força de cisalhamento das amostras de contrafilé aumentou (P < 0,01) com a inclusão de CZ na dieta, sendo esse resultado independente do tempo de maturação (7, 14, 21 ou 28 dias post-mortem). Observou-se interação entre período de administração de CZ e tempo de maturação (14 ou 28 dias post-mortem) para escore de maciez inicial (P = 0,03) e escore de tecido conjuntivo (P < 0,01), onde o processo de maturação melhorou mais a carne de animais controle do que a carne de animais suplementados com CZ. Mesmo com a diminuição da maciez, os bifes de animais suplementados com CZ foram considerados macios pelos provadores do painel sensorial. O CZ melhora a eficiência de produção de carne de bovinos Nelore, proporcionando aumento na massa muscular dos animais tratados sem alterar o consumo de matéria seca. Por outro lado, o tratamento dos animais com CZ reduz a maciez e a velocidade do processo de amaciamento do contrafilé. / The objective of this research was to evaluate the effects of zilpaterol hydrochloride (ZH) feeding time and feedlot days on performance, carcass characteristics, yield commercial cuts and meat quality of Nellore bulls. Ninety six Nellore bulls with initial body weight of 377 ± 25.1 kg were used in a randomized complete block in 4 × 2 factorial design, with four periods of ZH feeding (0, 20, 30 or 40 d) and 90 or 117 d in the feedlot. The supplementation period with ZH (8.33 mg/kg of dietary dry matter basis) started 23, 33 or 43 d before slaughter. In the last three days prior to slaughter all the animals were fed the diet without ZH, respecting the product grace period. Strip loin (m. Longissimus thoracis et lumborum) of all left-carcases were sampled for meat quality analysis. With the exception of LM area obtained both by ultrasound (P = 0.02) and by refrigerated carcasses (P = 0.05), there was no interaction between periods ZH feeding and days in the feedlot (P > 0.10). The increase of the supplementation period with ZH linearly increased feed efficiency (P < 0.01), hot carcass weight (P < 0.01), hot carcass yield (P < 0.01) and boning yield (P < 0.01). Beef strip loin steaks from animals supplemented with ZH presented a higher percentage of protein content (P < 0.01), while the 12th-rib fat (P = 0.04) and the percentage content of intramuscular fat (P < 0.01) decreased with ZH feeding. The percentage of polyunsaturated fatty acids increased (P = 0.01) with the ZH feeding and was mainly attributed to the increase of linoleic acid (18:2 cis-9, cis-12; P = 0.01) and arachidonic acid (20:4 n-6, P < 0.01). The Warner-Bratzler shear force (P < 0.01) from the strip loin samples increased with the inclusion of dietary ZH and this result was independent of the aging time (7, 14, 21 or 28 d postmortem). There was interaction between ZH feeding period and maturation time (14 or 28 d postmortem) to initial tenderness score (P = 0.03) and connective tissue score (P < 0.01), showing that the maturation process improved meat tenderness of control animals rather than ZH animals. Despite the initial meat tenderness decrease, strip loin steaks of animals supplemented with ZH were still considered tender by trained evaluators. ZH improves Nellore bulls meat production efficiency, providing an increase in muscle mass of the treated animals with no change in dry matter intake. However, treatment of animals with ZH reduces the tenderness and the speed of the strip loin maturation process.

and keywords The aim of this work was to evaluate the effect on rat myocardial β-adrenergic system of short-term (10 hours and 3 days) and long-term (5 weeks) cold exposure of rats with possible subsequent two-week recovery at normal temperature. The subtypes of β-adrenergic receptors (β-AR), their cognate G-proteins and adenylate cyclase have been characterized. β-AR are important components of cardiac regulatory mechanisms. They are involved in stimulating G-protein (Gs) and adenylate cyclase to increase cardiac contractility and frequency during stressful situations, including cold exposure. Heart tissue contains all three β-AR subtypes (β1-AR, β2-AR, β3-AR). While β1-AR couples with only Gs, β2-AR and β3-AR interact with the inhibitory G-protein (Gi). Electrophoresis and Western blotting showed a significant increase in β1-AR after a three-day exposure to cold. There was also a significant increase in β3-AR concentrations after a five-week cold exposure and this increase lasted for two weeks. There were no significant changes in the amnounts of β-AR cognate G-proteins (Gαs, Gβ, Gαi1/2 a Gαi3). In contrast, expression of adenylyl cyclase isoform V and VI significantly decreased during short-term exposure to cold. Using the saturation experiment with the β-antagonist [3H]CGP 12177, β-AR were...

Cardiovascular disease (CVD) is the leading cause of death in the United States. A common feature in most cardiac pathologies is the dysregulation of beta-adrenergic receptors (β-AR) and changes in the extracellular matrix (ECM). The ECM maintains strength and normal organization of cardiac tissue, while fibrosis (connective tissue scarring) is necessary for repair of damaged cardiac tissue. However, the dysregulation of the ECM leads to a number of cardiac disease pathologies. Osteopontin (OPN) is a protein with diverse biological functions in regulating the ECM such as bone resorption and calcification, wound healing, cell adhesion, cell survival, and apoptosis. OPN is expressed at low levels in the heart but increases with injury by promoting collagen synthesis, cardiac fibroblast growth, and adhesions to ECM proteins. Furthermore, as the heart ages, increases in ECM reorganization leads to cardiac damage and failure. Several studies have examined the role of OPN in the heart, but to date, no studies exist on the role of OPN in response to β-AR signaling and cardiac remodeling or the role that aging plays in this response. The goal of this study was to examine the effects of OPN on cardiac ECM remodeling following chronic beta-adrenergic stimulation. We proposed that OPN expression increases cardiac remodeling and dysfunction following ISO treatment in the aging heart evidenced by increased fibrosis. For this study, young (4 months) and middle age (14 months) mice with (WT) and without (KO) the OPN gene were treated with isoproterenol (ISO) for 28 days. Echocardiography was used to assess cardiac function. Mice were euthanized, and the hearts were analyzed for fibrosis using Masson’s Trichrome Staining. Results showed ISO increased fibrosis in the WT-ISO, but not KO-ISO compared to the respective controls (SHAM, no ISO treatment) for the middle age mice (p≤0.05). Furthermore, the aged WT-ISO group exhibited a 3-fold increase in fibrosis compared to the young WT-ISO group. Results from echocardiography will be analyzed and we expect to see compromised cardiac function in the WT-ISO groups compared to KO-ISO groups. OPN is currently being examined as a potential biomarker in heart failure. The results from this study will provide new insight on changes in the cardiac vasculature in the aging heart following injury and the role OPN plays in this process.

京都大学 / 0048 / 新制・論文博士 / 博士(医学) / 乙第13126号 / 論医博第2135号 / 新制||医||1024(附属図書館) / (主査)教授 湊谷 謙司, 教授 椛島 健治, 教授 岩井 一宏 / 学位規則第4条第2項該当 / Doctor of Medical Science / Kyoto University / DFAM

Indiana University-Purdue University Indianapolis (IUPUI) / Congenital heart defects occur in approximately one percent of births every year, which makes it the most frequently occurring congenital defect in patients. The aim of this project was to use two mutant neural crest (NC) mouse models to study the mechanisms underlying congenital heart failure in utero. The first mouse model was a Pax3 systemic knockout, which was lethal by mouse gestational day 14, and had appreciably reduced numbers of migratory NC cells. The second mouse model was a Wnt1Cre-mediated NC genetic cell ablation model, which was surprisingly viable and survived to birth, despite an apparent lack of migratory NC cells. The resultant data indicated that both mouse models had similar heart structural defects including persistent truncus arteriosus, which was due to fewer or no migratory cardiac NC cells. However, in utero heart function was appreciably perturbed in Pax3 mutants when compared to that of the ablated mutant model. The loss of embryonic cardiac function in Pax3 mutants was directly attributed to a substantial decrease in the activity of the beta-adrenergic pathway. This was due to a lack of proper specification of trunk NC cells, leading to diminished levels of circulating catecholamine levels in the embryo. To definitively confirm this conclusion, poor cardiac function was successfully restored by pharmacological stimulation of the beta-adrenergic pathway via administration of isoproterenol and forskolin to pregnant dams, which led to embryonic survival of Pax3 mutants to birth. By comparison of these two mutant mouse models, perturbation in the beta-adrenergic pathway was identified as the underlying mechanism responsible for in utero heart failure and lethality in Pax3 mutant embryos. The results of this study are expected to be significant in developing future therapeutic targets for congenital heart failure in prenatal and newborn patients.

No description available.

Thesis (MSc)--Stellenbosch University, 2004. / ENGLISH ABSTRACT: Please see fulltext for abstract. / AFRIKAANSE OPSOMMING: Sien asb volteks vir opsomming.

Thesis (PhD)--University of Stellenbosch, 2011. / ENGLISH ABSTRACT: The Mechanism of -adrenergic preconditioning ( -PC) Ischaemic preconditioning (IPC), a potent endogenous protective intervention against myocardial ischaemia, is induced by exposure of the heart to repetitive short episodes of ischaemia and reperfusion. The protective effects of this phenomenon have been demonstrated to be mediated by release of autocoids such as adenosine, opioids and bradykinin. Release of endogenous catecholamines and activation of the beta-adrenergic receptors (b-AR) have also been shown to be involved in ischaemic preconditioning. However, the exact mechanism whereby activation of the - adrenergic signal transduction pathway leads to cardioprotection, is still unknown. In view of the above, the aims of the present study were to evaluate: (i) the respective roles of the 1-, 2- and 3-AR receptors as well as the contribution of Gi protein and PKA to -adrenergic preconditioning, (ii) the role of the prosurvival kinases, PKB/Akt and ERK 44/p42 MAPKinase in -drenergic preconditioning, (iii) whether b-AR stimulation protect via ischaemia and the formation of adenosine; the respective roles of the A1-, A2-, A3-adenosine receptors as well as the involvement of the PI3-K/PKB/Akt and ERKp44/p42 signal transduction pathways, in the cardioprotective phenomemon of -adrenergic preconditioning and (iv) the contribution of the mitochondrial KATP channels (mKATP), reactive oxygen species and NO to the mechanism of -AR-induced cardioprotection. Methods: Isolated perfused rat hearts were subjected to 35 min regional ischaemia (RI) and reperfusion. Infarct size (IS) was determined using tetrazolium staining (TTC) and data were analyzed with ANOVA. Hearts were preconditioned with 5 min isoproterenol 0.1 μM ( 1/ 2-AR agonist), or formoterol 1 nM ( 2-AR agonist) or BRL 37344 1 μM ( 3-AR agonist) followed by 5 min reperfusion. The roles of the 1-, 2- and 3-ARs as well as NO were explored by using the selective antagonists CGP-20712A (300 nM), ICI -18551 (50 nM), SR59230A (100 nM) and NOS inhibitors L-NAME (50 μM) or LNNA (50 μM) respectively. Involvement of ROS and the mK+ ATP channels was studied by administration of N-acetyl cysteine (NAC, 300 μM) and the mitK+ ATP iv channel blocker 5-HD (100 μM) during the triggering phase. The role of PKA and PI3-K/Akt was investigated by the administration of the blockers Rp-8-CPT-cAMPs (16 μM) and wortmannin (100 nM) respectively, prior to RI or at the onset of reperfusion. Pertussis toxin (PTX), 30 μg kg-1 was administered i.p., 48 h prior to experimentation. The role of adenosine and the adenosine A1, A3, A2A and A2B receptors was studied by using adenosine deaminase and the selective antagonists DPCPX (1 μM), MRS 1191(1 μM), ZM241385 (1 μM) and MRS1754 (1 μM). Activation of PKB/Akt and ERKp44/p42 was determined by Western blot. Results: Infarct sizes of hearts preconditioned with isoproterenol of formoterol were significantly smaller compared to those of non-preconditioned hearts. This was associated with an improvement in postischaemic mechanical performance. However the 3-AR agonist BRL37344 could not reduce infarct size. The 1- and 2-AR blockers CGP-20712A and ICI-118551 completely abolished the isoproterenol-induced reduction in infarct size and improvement in mechanical recovery, while the 3-AR blocker was without effect. Both Rp-8-CPT-cAMPs and wortmannin significantly increased infarct size when administered before 1/ 2-AR preconditioning or at the onset of reperfusion while it reduced mechanical recovery during reperfusion. PTX pretreatment had no significant effect on the reduction in infarct size induced by 1/ 2-AR or 2-AR preconditioning, however it reduced mechanical recovery in the latter. The NOS inhibitors had no effect on the reduction in infarct size induced by 1/ 2-AR preconditioning, but depressed mechanical function during reperfusion. The significant reduction in infarct size by 1/ 2-PC, was associated with activation of ERKp44/p42 and PKB/Akt during the triggering phase, as well as during reperfusion. DPCPX (A1-AdoR antagonist) had no effect on the 1/ 2-PC-induced reduced infarct size or ERK p44/p42 and PKB activation. A2A-AdoR, but not A2b-AdoR, blockade during the trigger phase abolished the reduction in infarct size of 1/ 2-PC. Both antagonists significantly reduced ERK and PKB activation in the trigger phase. In addition, when applied at the onset of reperfusion they significantly reduced ERK p44 / v p42 MAPK and PKB/Akt activation to an even greater extent. MRS-1191 (A3-AdoR antagonist) blocked 1/ 2-PC when applied prior to index ischaemia or when added during early reperfusion, significantly inhibiting both ERK p44 and PKB activation. Cardioprotection of 1/ 2-PC was abolished by inhibition of ROS generation with NAC in the triggering phase as well as at the start of reperfusion. However, the mitoK+ ATP channel blocker 5- HD was without effect. Conclusions: Protection afforded by an acute transient stimulation of the -ARs, depends on the activation of both 1-AR and 2-ARs but not the 3-AR. PKA as well as PI3-K activation prior to sustained ischemia and at the onset of reperfusion were essential for cardioprotection. With functional recovery as endpoint, it appears that NO is involved in 1/ 2-AR preconditioning, while the Gi protein may play a role in 2-AR preconditioning. The production of endogenous adenosine induced by transient b1/b2 stimulation of the isolated rat heart is involved in b−AR preconditioning. Cardioprotection was shown not to be dependent on the A1AdoR while activation of the A3-AdoR occurs during both the triggering and mediation phases. Both the adenosine A2A and, to a lesser extent, the adenosine A2B receptors participate in the triggering phase of b1/b2-PC. Generation of ROS during the triggering and reperfusion phases is involved in eliciting protection, but no role for the mKATP channels could be demonstrated. Finally, activation of the RISK pathway (PKB/Akt and ERKp44/p42) during the triggering phase is a prerequisite for protection. In addition, cardioprotection by b-AR is characterized by activation of the RISK pathway during reperfusion. / AFRIKAANSE OPSOMMING: Iskemiese prekondisionering (IPC) is ‘n kragtige endogene beskerming teen miokardiale iskemie, wat deur blootstelling van die hart aan kort opeenvolgende episodes van iskemie en herperfusie, ontlok word. Hierdie beskerming word medieer deur vrystelling van outakoïede soos adenosine, opioïede en bradikinien. Vrystelling van endogene katekolamiene en aktivering van die betaadrenerge reseptore (b-AR) is bewys om ook by hierdie proses betrokke te wees. Die presiese meganismes waardeur aktivering van die -adrenerge seintransduksiepad tot miokardiale beskerming lei, is nog onbekend. In die lig van bogenoemde, was die doel van die huidige studie om die volgende te evalueer: (i) die onderskeie rolle van die b1-, b2- en b3-AR sowel as die bydrae van die Gi proteïen en PKA in b- adrenerge prekondisionering, (ii) of b-AR stimulasie beskerming ontlok via iskemie en vorming van adenosien, die onderskeie rolle van die A1-, A2-, A3-adenosien reseptore (AdoRs) sowel as die PI3- K/PKB/Akt en ERKp44/p42 seintransduksie paaie, (iv) die mitochondriale KATP (mKATP) kanale, vry suurstof radikale en NO in b−AR prekondisionering. Metodes: Geïsoleerde, geperfuseerde rotharte is aan 35 minute streeksiskemie en herperfusie onderwerp. Infarktgrootte (IS) is deur die tetrazolium (TTC)-kleuringsmetode bepaal. Data is met behulp van ANOVA analiseer. Harte is geprekondisioneer vir 5 min met isoproterenol 0.1 μM ( 1/ 2-AR agonist), of formoterol 1 nM ( 2-AR agonist) of BRL 37344 1 μM ( 3-AR agonist), gevolg deur 5 min herperfusie, voor streeksiskemie. Die belang van die 1-, 2- en 3-ARs sowel as NO is ondersoek, deur onderskeidelik gebruik te maak van selektiewe antagoniste nl CGP- 20712A (300 nM), ICI -18551 (50 nM), SR59230A (100 nM) en NOS inhibitore L-NAME (50μM) of LNNA (50μM). Die rol van die mK+ ATP kanale en ROS is bepaal deur die toediening van die mK+ ATP kanaal blokker 5-HD (100 μM) en die vrye-radikaal opruimer, N-asetiel cysteine (NAC, 300 μM). Die belang van PKA en PI3-K/Akt is bepaal deur toediening van die PKA blokker Rp-8- CPT-cAMPs (16μM) en wortmannin (100nM) respektiewelik. Pertussis toxin (PTX), 30 μg kg-1 is i.p toegedien, 48 uur voor eksperimentasie. vii Die rol van adenosien en die adenosien A1, A2A, A2B en A3 reseptore is bestudeer, deur gebruik te maak van adenosien deaminase en die selektiewe antagoniste DPCPX (1 μM), MRS 1191(1 μM), ZM241385 (1 μM) and MRS1754 (1 μM),repektiewelik. Die middels is deurgaans toegedien tydens die prekondisioneringsprotokol (“snellerfase”) of tydens vroeë herperfusie. Aktivering van PKB/Akt en ERK p44/p42 is deur Western blot analise bepaal. Resultate: Infarktgrootte van harte wat geprekondisioneer is met of isoproterenol ( 1/ 2-PC) of formoterol ( 2-PC), was beduidend kleiner as díe van ongeprekondisioneerde harte. Dit is geassosieer met ‘n toename in postiskemiese meganiese herstel. Die 3-AR agonis BRL37344 ( 3- PC) het egter geen effek op infarktgrootte gehad nie. Die selektiewe 1- en 2-AR blokkers CGP- 20712A en ICI-118551 het die afname in infarktgrootte heeltemal opgehef, asook die verbetering in meganiese herstel tydens herperfusie terwyl die 3-AR blokker geen effek getoon het nie. Beide Rp- 8-CPT-cAMPs en wortmannin het infarktgrootte beduidend vergroot en meganiese herstel beduidend verlaag, wanneer dit net voor 1/ 2-prekondisionering of tydens die begin van herperfusie toegedien is. PTX voorafbehandeling het geen beduidende effek op die vermindering van infarktgrootte (geïnduseer deur 1/ 2-PC of 2-PC) gehad nie. Meganiese herstel is egter verminder in die geval van 2-PC. Die NOS inhibitore het geen effek op die vermindering in infarktgrootte geïnduseer deur b1/b2 gehad nie, maar het ook meganiese herstel onderdruk. Die beduidende afname in infarktgrootte deur b1/b2 prekondisionering is gekenmerk deur aktivering van ERKp42/p44 en PKB/Akt tydens die snellerfase. Soortgelyke aktivering van hierdie kinases is ook tydens herperfusie van b-AR geprekondisioneerde harte waargeneem. DPCPX (A1-AdoR antagonis) het geen effek op die infarkt-verminderde effek van 1/ 2- prekondisionering of op ERK p44/p42 en PKB aktivering gehad nie. A2A-AdoR, maar nie A2b – AdoR, blokkade tydens die snellerfase, het die effek van b-AR prekondisionering op infarktgroottee opgehef. Beide antagoniste het die aktivering van ERKp42/p44 en PKB/Akt tydens die snellerfase onderdruk. Wanneer toegedien tydens herperfusie, het dit die aktivering van hierdie kinases tot ‘n groter mate onderdruk. MRS-1191 (A3-AdoR antagonis) het infarktgrootte beduidend verhoog en 1/ 2-prekondisionering geblokkeer, beide wanneer dit voor indeks-iskemie toegedien is of tydens vroeë herperfusie, tesame met inhibisie van PKB en ERK p44/p44 aktivering. viii Die kardiobeskerming van 1/ 2-prekondisionering is opgehef deur middel van opruiming van vry suurstof radikale deur NAC in die snellerfase sowel as aan die begin van herperfusie. Die mK+ ATP kanaal blokker 5-HD het geen effek op b-AR prekondisionering gehad nie. Gevolgtrekking: Kardiobeskerming teweeggebring deur ‘n kort periode van stimulasie van die - ARs, is afhanklik van die aktivering van beide 1-AR en 2-ARs, maar nie 3-AR nie. PKA sowel as PI3-K aktivering, net voor volgehoue iskemie en tydens vroeë herperfusie, is aangedui om noodsaaklik vir 1/ 2-AR prekondisionering te wees. Waar funksionele herstel as eindpunt gebruik is, blyk dit dat NO wel betrokke is by 1/ 2-AR prekondisionering, terwyl die Gi protein ‘n rol mag speel in 2-AR prekondisionering. Vorming van endogene adenosien tydens b-adrenerge stimulasie is betrokke by b-AR prekondisionering. Hierdie beskerming is nie van die A1-AdoR afhanklik nie, maar aktivering van die A3-AdoR is nodig tydens beide die sneller en herperfusie fases. Beide die A2A-AdoR, en tot ‘n mindere mate die A2B–AdoR, is ook betrokke by die snellerfase. Vorming van vry suurstof radikale is nodig vir b-AR prekondisionering, nterwyl die mKATP kanale nie betrokke is nie. Ten slotte, aktivering van die RISK seintransduksiepad (ERKp42/p44 en PKB/Akt) tydens die snellerfase is ‘n voorvereiste vir die ontlokking van beskerming. Daarbenewens word b-AR prekondisionering gekarakteriseer deur aktivering van hierdie pad tydens herperfusie. / South African Medical Research Council / University of Stellenbosch