## 4 Discussion  
This study had seven principal findings. (1) mRNA gene expression of BCL2 was significantly increased (\(p<0.01\)) by exposure for one hour to selective adenosine A2A receptor agonist CGS21680. (2) mRNA gene expression of BCL2 was not significantly affected by exposure for one hour to adenosine analogue NECA. (3) mRNA gene expression of BCL2 was not significantly affected by selective adenosine A2B receptor antagonist, PSB 603 (100 nM or 1 uM). (4) Baseline BCL2 mRNA gene expression was significantly reduced by the addition of NECA and 100 nM (but not 1 uM) of PSB 603 when the PSB 603 was added to cells ten minutes before the NECA was added. (5) Expression of BCL2 was not significantly affected by Forskolin (100 nM). (6) Expression of BCL2 was not modulated by insulin (100 nM). (7) In skeletal muscle cells, the action of insulin was not modulated by NECA. Consistent with the original hypothesis of this study, the findings in the study for the first time showed that after CGS21680 treatment, BCL2 gene expression increased inside skeletal muscle cells, exposing them to a possible anti-apoptotic effect. Therefore, the potential increase in BCL2 protein expression could play a protective role against apoptosis, preventing skeletal muscle muscular damage [8]. In fact, the results showed that the adenosine A2A receptor's potential stimulation played a role in modulating the expression of the BCL2 gene that appears during CGS21680 treatment. The results also showed the adenosine A2A receptor agonists to be potentially useful drugs in the treatment of skeletal muscle injury. Adenosine A2A receptor agonists were potentially important apoptosis regulators [9; 13]. Based on these results, it was therefore suggested that such adenosine A2A receptor agonists as CGS 21680 might be effective in treating a number of skeletal muscle regeneration diseases as well [9; 23]. This study investigated the selective adenosine A2A receptor agonists' involvement in apoptosis modulation. The  
Figure 4: Effects of insulin and/or NECA on BCL2 messenger RNA (mRNA) gene expression, using charcoal serum, in rat L6 skeletal muscle myotubes. Data were analyzed with a one-way ANOVA test, then a Tukey test. current investigation therefore introduced adenosine A2A agonists as potential agents for muscular dystrophy pharmacotherapy. Further research is required to achieve a complete understanding of apoptosis' molecular basis relating to adenosine receptors. This study begins to explain the mechanism by which adenosine A2A receptor agonist CGS21680 can prevent skeletal muscle apoptosis. Therefore, the receptors of adenosine could also represent novel pathways or targets for the pharmacological intervention in the process of apoptosis. CGS21680, at the concentration applied in this current study, was regarded as a selective adenosine A2A receptor agonist (Ki = 27 nM) [24]. However, some uncertainty surrounds the efficacy of CGS21680 targets due to the differences in various species' binding affinity of ligands for adenosine receptors and the lack of binding studies performed on rats. Of note, PSB 603 showed a strong affinity to adenosine A2B receptors but only a very weak affinity to three other subtypes of adenosine receptor. Adenosine A2B receptors show >17,000 times the selectivity of other adenosine receptors (Ki values: 0.553, >10,000, >10,000, and >10,000 nM for A2B, A1, A2A and A3 receptors, respectively) [25]. Experiments were conducted to investigate whether the adenylyl cyclase pathway contributes to the activation, in skeletal muscle cells, of the BCL2 transcription profile. Because of this, the present study used the adenylyl cyclase activator forskolin to help discover if the cAMP pathway plays a potential role in NECA-induced BCL2 mRNA gene expression in skeletal muscle. Previous studies showed that activation, by NECA, of adenosine A2B receptors in skeletal muscle increased the NR4A mRNA gene expression [19] and cAMP accumulation [14].