AIMTo evaluate the effects of beta3-adrenergic receptors (ARs) agonist (BRL-37344) on beating rate and cAMP levels and investigate the influence on the chronotropic action of beta3-ARs in cultured cardiomyocyte of rats.

METHODSCultured neonatal rat cardiomyocytes were divided randomly into eight groups, control group, ISO group, Nadolol + ISO group, BRL group, PIX + BRL group, L-NAME + BRL group, Nadolol + BRL group and Bupranolol + BRL group. Beating rate of culture neonatal rat cardiomyocytes was observed and cAMP measured by enzyme immunoassay kit. Expression levels of beta3-ARs mRNA in cardiomyocytes was evaluated by reverse transcription-polymerase chain reaction (RT-PCR).

RESULTSISO, nonspecific beta-ARs agonist increased beating rate and intracellular cAMP production, antagonized by Nadolol, beta1, beta2-ARs antagonist. BRL37344 decreased beating rate and intracellular cAMP levels. FPTX, Gi protein inhibitor and Bupranolol, nonspecific beta-ARs antagonist totally blocked the effect and L-NAME, nitric oxide synthase (NOS) inhibitor partly blocked the effect, but Nadolol did not. There was the expression of beta3-AR mRNA in cardiomyocytes by RT-PCR.

CONCLUSIONSBeta3-ARs showed in cardiomyocytes and produced negative chronotropic effects. beta1, beta2-ARs antagonist did not affect it. It suggested beta3-ARs signal transduction was related with G1 protein. The negative inotropic effect of beta3-ARs stimulation was mediated by activation of the NOS pathway.

Adrenergic beta-Agonists ; pharmacology ; Animals ; Animals, Newborn ; Cells, Cultured ; Cyclic AMP ; metabolism ; Ethanolamines ; pharmacology ; Myocytes, Cardiac ; drug effects ; metabolism ; RNA, Messenger ; genetics ; Rats ; Rats, Wistar ; Receptors, Adrenergic, beta-3 ; metabolism

OBJECTIVETo observe the effect of beta(3)-adrenoceptor (AR) in regulating resting intracellular Ca(2+) concentration of the ventricular myocytes and investigate the signaling pathway in rats with experimental heart failure.

METHODSRat models of experimental heart failure were established by ligation of the anterior descending artery, and the myocytes were isolated by enzymatic digestion. The resting intracellular Ca(2+) concentration was determined using laser scanning confocal microscopy (LSCM) in the cells stimulated with 1 micromol/L BRL37344 (a selective beta(3)-AR agonist) alone or in combination with PTX, L-NAME, or methylene blue.

RESULTSIn the ventricular myocytes from normal control rats, BRL373444 reduced the resting intracellular Ca(2+) concentration of by 45.5%, while the reduction increased to 59.4% in the cells from rats with heart failure. In combination with L-NAME (10 micromol/L), methylene blue (10 micromol/L), and PTX (2 microg/ml), BRL373444 caused a reduction in resting intracellular Ca(2+) concentration of the ventricle myocytes from normal control rats by 10.1%, 16.9%, and 15.4%, respectively in control group, while the rate was 16.9%, 19.3%, and 11.7% in the heart failure group.

CONCLUSIONSBeta(3)-AR agonist can decrease the resting intracellular Ca(2+) concentration of the ventricular myocytes, but the reduction is smaller in cells from rats with heart failure than in cells of normal rats. This effect is mediated through the PTX-NOS-NO pathway.

Adrenergic Agonists ; pharmacology ; Adrenergic beta-3 Receptor Agonists ; Animals ; Calcium ; metabolism ; Heart Failure ; chemically induced ; metabolism ; pathology ; Heart Ventricles ; pathology ; In Vitro Techniques ; Intracellular Space ; drug effects ; metabolism ; Male ; Myocytes, Cardiac ; drug effects ; metabolism ; pathology ; Rats ; Rats, Wistar ; Receptors, Adrenergic, beta-3 ; metabolism ; Rest ; Signal Transduction ; drug effects

OBJECTIVETo evaluate the effects of different doses of N(G)-nitro-L-arginine methyl ester (L-NAME) on hemodynamics, cyclic guanosine monophosphate (cGMP) production and the level of beta-adrenergic receptors (beta-ARs) mRNA in a heart failure rat model after BRL-37344 (beta(3)-ARs agonist) injection. Meanwhile, to investigate the influence of beta(3)-ARs and L-NAME on signal transduction in failing heart.

METHODSThe rats were randomly divided into six groups, control group (group I), Iso (isoproterenol) group (group II), Iso + BRL group (group III), Iso + BRL + low dose of L-NAME group (5 mg/kg, group IV), Iso + BRL + moderate dose of L-NAME group (50 mg/kg, group V), Iso + BRL + high dose of L-NAME group (100 mg/kg, group VI). The hemodynamics [left ventricular end systolic pressure (LVESP), +/- dp/dt, left ventricular end diastolic pressure (LVEDP)], cardiac cGMP and the levels of beta(1)-, beta(2)-, and beta(3)-ARs mRNA were measured.

RESULTS(1) LVESP, +/- dp/dt values in group II were significantly lower, and LVEDP was significantly higher than that in group I (except -dp/dt P < 0.05, the rest were P < 0.01). Comparing with group II, group III had lower -dp/dt value and LVESP, higher LVEDP (P < 0.05). The level of +dp/dt had a trend to be lower but lacked statistical significance between two groups. The value of +/- dp/dt got higher and LVEDP got lower along with higher dose of L-NAME, but a large dose of L-NAME had more deteriorated cardiac functions. (2) The cardiac cGMP in group I, II and III had a higher tendency (P < 0.01). The tendency of cardiac cGMP in group IV, V and VI was inversed with the dose of L-NAME. After a large dose of L-NAME was applied, cGMP returned to the same level as Group I. (3) Among groups I, II and III, the level of beta(1)-AR mRNA was the highest in group I and the lowest in group III (P < 0.01). The levels of beta(2)-AR mRNA were also tended to be lower among three groups but with no significance. While the level of beta(3)-AR mRNA was the highest in group III. The levels of beta-AR mRNA were all the same in group VI, V and VI.

CONCLUSIONSThe negative inotropic effect of beta(3)-ARs stimulation was mediated by activation of the NOS pathway. L-NAME blocked beta(3)-ARs agonist negative chronotropic effect on failing heart partly and improved hemodynamics, but a large dose of L-NAME had more deteriorated cardiac functions.

Adrenergic Agonists ; therapeutic use ; Animals ; Cyclic GMP ; metabolism ; Heart Failure ; drug therapy ; metabolism ; physiopathology ; Male ; NG-Nitroarginine Methyl Ester ; administration & dosage ; pharmacology ; RNA, Messenger ; metabolism ; Rats ; Rats, Wistar ; Receptors, Adrenergic, beta-3 ; metabolism

The effects of glucagon and epinephrine on gluconeogenesis in young (4 month) and old (24 month) Fisher 344 rat hepatocytes were compared. In contrast to glucagon, which had a similar effect on gluconeogenesis in both young and old cells, epinephrine caused a smaller increase in gluconeogenesis in old rat hepatocytes than in young hepatocytes. beta2 adrenergic receptor (beta2-AR) expression slightly decreased in aged rat liver, and there were differences between young and old hepatocytes in their patterns of G protein coupled receptor kinases, which are involved in the activation of beta2-AR receptor signal desensitization. The major isoform of the kinase changed from GRK2 to GRK3 and the expression of beta-arrestin, which is recruited by the phosphorylated beta2-AR for internalization and degradation, increased in aged rat liver. GRK3 overexpression also decreased the glucose output from young rat hepatocytes. We conclude that an age-associated reduction in epinephrine-induced gluconeogenesis occurs through the epinephrine receptor desensitizing system.
Adrenergic beta-Agonists/*pharmacology ; Aging/*drug effects ; Animals ; Epinephrine/*pharmacology ; G-Protein-Coupled Receptor Kinase 2/metabolism ; G-Protein-Coupled Receptor Kinase 3/metabolism ; Glucagon/pharmacology ; *Gluconeogenesis/drug effects ; Male ; Models, Biological ; Phosphorylation ; Rats ; Rats, Inbred F344 ; Receptors, Adrenergic, beta-2/agonists/metabolism

The effects of glucagon and epinephrine on gluconeogenesis in young (4 month) and old (24 month) Fisher 344 rat hepatocytes were compared. In contrast to glucagon, which had a similar effect on gluconeogenesis in both young and old cells, epinephrine caused a smaller increase in gluconeogenesis in old rat hepatocytes than in young hepatocytes. beta2 adrenergic receptor (beta2-AR) expression slightly decreased in aged rat liver, and there were differences between young and old hepatocytes in their patterns of G protein coupled receptor kinases, which are involved in the activation of beta2-AR receptor signal desensitization. The major isoform of the kinase changed from GRK2 to GRK3 and the expression of beta-arrestin, which is recruited by the phosphorylated beta2-AR for internalization and degradation, increased in aged rat liver. GRK3 overexpression also decreased the glucose output from young rat hepatocytes. We conclude that an age-associated reduction in epinephrine-induced gluconeogenesis occurs through the epinephrine receptor desensitizing system.
Adrenergic beta-Agonists/*pharmacology ; Aging/*drug effects ; Animals ; Epinephrine/*pharmacology ; G-Protein-Coupled Receptor Kinase 2/metabolism ; G-Protein-Coupled Receptor Kinase 3/metabolism ; Glucagon/pharmacology ; *Gluconeogenesis/drug effects ; Male ; Models, Biological ; Phosphorylation ; Rats ; Rats, Inbred F344 ; Receptors, Adrenergic, beta-2/agonists/metabolism

&beta;3-adrenoceptor (&beta;3-AR) has been shown to promote myocardial apoptosis. However, the exact physiological role and importance of this receptor in the human myocardium, and its underlying mode of action, have not been fully elucidated. The present study aimed to determine the effects of &beta;3-AR on the promotion of myocardial apoptosis and on norepinephrine (NE) injury. We analyzed NE-induced cardiomyocyte (CM) apoptosis by using a TUNEL and an annexin V/propidium iodide apoptosis assay. Furthermore, we investigated the NE-induced expression of the apoptosis marker genes Akt and p38MAPK, their phosphorylated counterparts p-Akt and p-p38MAPK, caspase-3, Bcl-2, and Bax. In addition, we determined the effect of a 48-h treatment with a &beta;3-AR agonist and antagonist on expression of these marker genes. &beta;3-AR overexpression was found to increase CM apoptosis, accompanied by an increased expression of caspase-3, bax/bcl-2, and p-p38MAPK. In contrast, the &beta;3-blocker reduced apoptosis of CMs and the associated elevated Akt expression. We identified a novel and potent anti-apoptosis mechanism via the PI3K/Akt pathway and a pro-apoptosis pathway mediated by p38MAPK.
Adrenergic Agonists ; pharmacology ; Adrenergic Antagonists ; pharmacology ; Animals ; Apoptosis ; Cells, Cultured ; Myocytes, Cardiac ; drug effects ; metabolism ; Phosphatidylinositol 3-Kinases ; metabolism ; Proto-Oncogene Proteins c-akt ; metabolism ; Rats ; Rats, Sprague-Dawley ; Receptors, Adrenergic, beta-3 ; genetics ; metabolism ; Signal Transduction ; p38 Mitogen-Activated Protein Kinases ; metabolism

We sought to determine the effects of activation of peroxisome proliferator-activated receptor-gamma (PPAR-gamma) on multilocularization of adipocytes in adult white adipose tissue (WAT). Male C57BL/6 normal, db/db, and ob/ob mice were treated with agonists of PPAR-gamma, PPAR-alpha, or beta3-adrenoceptor for 3 weeks. To distinguish multilocular adipocytes from unilocular adipocytes, whole-mounted adipose tissues were co-immunostained for perilipin and collagen IV. PPAR-gamma activation with rosiglitazone or pioglitazone induced a profound change of unilocular adipocytes into smaller, multilocular adipocytes in adult WAT in a time-dependent, dose-dependent, and reversible manner. PPAR-alpha activation with fenofibrate did not affect the number of locules or remodeling. db/db and ob/ob obese mice exhibited less multilocularization in response to PPAR-gamma activation compared to normal mice. Nevertheless, all adipocytes activated by PPAR-gamma contained a single nucleus regardless of locule number. Multilocular adipocytes induced by PPAR-gamma activation contained substantially increased mitochondrial content and enhanced expression of uncoupling protein-1, PPAR-gamma coactivator-1-alpha , and perilipin. Taken together, PPAR-gamma activation induces profound multilocularization and enhanced mitochondrial biogenesis in the adipocytes of adult WAT. These changes may affect the overall function of WAT.
Adipocytes/*cytology/metabolism ; Adipose Tissue, White/*cytology ; Animals ; Cell Nucleus Division ; Hypoglycemic Agents/pharmacology ; Ion Channels/metabolism ; Male ; Mice ; Mice, Inbred C57BL ; Mice, Obese ; Mitochondria/metabolism ; Mitochondrial Proteins/metabolism ; PPAR alpha/agonists/metabolism ; PPAR gamma/*agonists/*metabolism ; Phosphoproteins/metabolism ; Receptors, Adrenergic, beta-3/agonists ; Thiazolidinediones/pharmacology ; Trans-Activators/metabolism