**2. Objectives**

increase of insulin levels and lipolysis after administration of CL316243 in normal mice was noted; effects were not found after administration of beta-3 adrenergic agonists in beta-3

In another model of knockout mice lacking functional beta-3 adrenoreceptors, there were no responses for food intake and insulin secretion in white and brown adipocytes after administration of beta-3 adrenergic agonist CL316243, indicating the implication of beta-3 receptors in

An increase in insulin levels during "fasted/fed" transition in rats has been demonstrated, associated with a decrease in the mRNA level of beta-3 adrenergic receptor and a decrease of the response in brown and white adipose tissues. It was concluded that there is a close relationship between the food intake, plasma levels of insulin, and beta-3 adrenergic receptors. Downregulation of the beta-3 receptors could be a possible mechanism by which insulin

In another study, CL316243 was administered in obese diabetic KKAy mice for 2 weeks. The results showed a decrease of serum levels of glucose, insulin, triglyceride, free fatty acid, and tumor necrosis factor-alpha (TNF-alpha) and an increase of adiponectin. The beta-3 adrenergic receptor agonist recovered the mRNA expressions of adiponectin, adiponectin receptors, and beta-3 adrenoreceptor, which were reduced in epididymal white adipose tissue in KKAy mice. Also, CL316243 suppressed the overexpressed mRNA level of TNF-alpha in both epididymal white and brown adipose tissues. It was concluded that the normalization of adiponectin, adiponectin receptors and TNF-alpha could contribute at the amelioration of obesity-induced

In the study conducted on nonobese/nondiabetic Sprague-Dawley rats, the selective beta-3 agonist CL 312243 increased food intake, metabolic rate, and body temperature after 7 days of treatment. The author also showed a decrease in intra-abdominal and subepithelial fat, a hepatic glucose level independent of variations in body weight, an increase in interscapular fat, and in total glucose, which stimulates the production of insulin. According to the results, only white and brown adipose tissues have been affected. A more important role of adipose tissue in glucose uptake underlining the potential role of beta-3 adrenergic agonist drugs for

By a critical analysis of the published data in nonclinical research, it was concluded that beta-3 adrenergic receptor activation in the experiment-induced diabetes and obesity [37, 38] determines an increase of glucose tolerance and lipolysis activation in adipose tissue [22]. It seems that the effects are dose dependent and the selectivity of actions for glucose metabolism

Comparing the effects of beta-3 agonists in rats and humans, Arch and Wilson stated that these compounds, with remarkable effects on rodents, have not convinced in clinical studies because of limited efficacy or serious side effects. The explanations mentioned by the authors included low pharmacokinetic properties and a low biotransformation to active compounds. A possible more important distinction between rats and humans is the different structure of beta-3 receptors, leading to lower efficacy of compounds in humans than in rats. In addition, it seems

determines lipid storage and prevents lipid mobilization after food intake [36].

the treatment of obesity and insulin-resistant diabetes was suggested [8].

occurs at lower doses than those used for influencing lipid metabolism.

receptor-deficient mice [34].

224 Adiposity - Epidemiology and Treatment Modalities

these metabolic effects [35].

insulin resistance [28].

The purpose of this study was to test the effects of the newly synthesized compounds over lipid profile and body weight of rats to which alloxanic diabetes was induced, being a known fact that this metabolic disorder induces alterations in plasma lipids.

Alloxan was chosen for induction of type II diabetes mellitus because its pancreatic toxicity was demonstrated in nonclinical trials using isolated islet cell or entire perfused rat pancreas and afterwards by directly administering the substance to rodent or nonrodent animals. In the first stage, alloxan stimulates on short-term insulin secretion, which is followed by total suppression of the response of the islet cells to glucose, regardless of its concentration [43].

Alloxan is readily absorbed by beta pancreatic cell, a process that contributes to diabetogenic action. Its absorption also takes place in liver although hepatocytes are more resilient to its action compared to beta cells, therefore more protected against its toxicity [44].

#### **2.1. Mechanism of alloxan toxicity**

The mechanism of toxic action resides in formation of reactive species of oxygen [45], the alloxan exhibiting increased affinity to substrates containing thiolic groups (reduced glutathione, cysteine, proteins with sulfide groups, enzymes) [46]. The product of alloxan reduction, the dialuric acid, is reoxidized to alloxan, the redox cycle thus formed being responsible for releasing superoxide radicals—see **Figure 2** [47].

**Figure 2.** Mechanism of diabetes induction by administration of alloxan [47]. A, Alloxan; AH·, alloxan radical; AH2, dialuric acid; O2· - , superoxide radical; OH·, hydroxyl radical.

An optimal protection against cytotoxic action of alloxan and dialuric acid is offered by an association between superoxide-dismutase (SOD) and catalase (CAT), in order to completely prevent the redox cycle and consecutively the formation of any reactive oxygen species [48].

Glucose confers complete protection against toxic effects of alloxan both *in vivo* and *in vitro*, by blocking glucokinase inhibition by it and also contributing to maintaining the antioxidant protection mechanism of the beta cells [49].

One of the targets for the reactive oxygen species is the DNA of the pancreatic islet cells. The DNA fragmentation occurs in beta cells exposed to alloxan [49, 50].

The DNA alteration stimulates poly-ADP-ribose, enzyme which contributes to affected DNA repair. In several trials it was stated that glucose administration contributes to counteracting the alloxan cytotoxicity. Such ability is the result not only of glucokinase protection but also of interaction with glucose carrier GLUT2 resulting in reduced alloxan absorption [51].
