To test the hypothesis that the high circulating FFA levels in the diabetes of obesity could contribute to the altered dynamics of insulin secretion seen in that condition, insulin release was measured in isolated perifused rat islet cells, without or with added palmitate. Acutely, as in other systems, palmitate (1 mM) stimulated insulin release. Palmitate (1 mM) suppressed both first and second phase insulin release after 2, 3, or 4 h of perifusion, but not after 1 h. No significant effect was noted with 0.3 mM palmitate, and the effect was maximal at 1 mM. The stimulatory effects of arginine were essentially unaffected. Tolbutamide (1 mM) reversed or counteracted the effect. Glucose oxidation was suppressed in islets incubated with 1 mM palmitate for 4 h. Inhibitors of fat oxidation, alpha-bromostearate (1 mM) and methyl-3-tetradecylglycidate (100 microM) reversed the effects of palmitate on glucose-stimulated insulin release and glucose oxidation. Thus, prolonged incubation of rat islet cells with 1 mM palmitate could suppress the glucose-stimulated release of insulin from perifused rat islets. This suppression could be reversed by inhibitors of fat oxidation. This supports the hypothesis that elevated FFA levels and/or increased fat oxidation could contribute to the altered dynamics of insulin secretion in obese diabetics by fuel antagonism as well as the previously documented suppression of peripheral glucose uptake and stimulation of hepatic gluconeogenesis and may be a key link between obesity and the development of diabetes.

Correlations (Corrected and Observed) of New Medical College Admission Test (MCAT) Total Scores and Undergraduate Grade Point Averages (UGPAs), Alone and Together, with Students' Summative Performance a in the First Year of Medical School (M1) Data type Corrected b (observed) correlations of MCAT total scores with summative performance in M1 Corrected (observed) correlations of UGPAs with summative performance in M1 Corrected (observed) correlations of MCAT total scores and UGPAs with summative performance in M1 Median 0.57 (0.39) 0.52 (0.34) 0.65 (0.52) Interquartile range (25%-75%)

To test the hypothesis that the high circulating FFA levels in the diabetes of obesity could contribute to the altered dynamics of insulin secretion seen in that condition, insulin release was measured in isolated perifused rat islet cells, without or with added palmitate. Acutely, as in other systems, palmitate (1 mM) stimulated insulin release. Palmitate (1 mM) suppressed both first and second phase insulin release after 2, 3, or 4 h of perifusion, but not after 1 h. No significant effect was noted with 0.3 mM palmitate, and the effect was maximal at 1 mM. The stimulatory effects of arginine were essentially unaffected. Tolbutamide (1 mM) reversed or counteracted the effect. Glucose oxidation was suppressed in islets incubated with 1 mM palmitate for 4 h. Inhibitors of fat oxidation, alpha-bromostearate (1 mM) and methyl-3-tetradecylglycidate (100 microM) reversed the effects of palmitate on glucose-stimulated insulin release and glucose oxidation. Thus, prolonged incubation of rat islet cells with 1 mM palmitate could suppress the glucose-stimulated release of insulin from perifused rat islets. This suppression could be reversed by inhibitors of fat oxidation. This supports the hypothesis that elevated FFA levels and/or increased fat oxidation could contribute to the altered dynamics of insulin secretion in obese diabetics by fuel antagonism as well as the previously documented suppression of peripheral glucose uptake and stimulation of hepatic gluconeogenesis and may be a key link between obesity and the development of diabetes.

Exposure of 3T3-L1 adipocytes to 1 nM insulin for 10 min results in activation of particulate cAMP phosphodiesterase and suppression of lipolysis stimulated by 10 nM isoproterenol. When lipolysis was increased by cilostamide, a selective inhibitor of the particulate phosphodiesterase, the antilipolytic effect of insulin was not observed. Insulin did suppress lipolysis stimulated by Ro 20-1724, an inhibitor of soluble cAMP phosphodiesterase activity. Cilostamide did not interfere with insulin stimulation of glucose uptake, nor did it have any direct effect on cAMP-dependent protein kinase. Thus, inhibition of particulate but not soluble cAMP phosphodiesterase blocked the antilipolytic effect of insulin. Our findings support the idea that insulin inhibits lipolysis, perhaps in large part by activating particulate "low Km" cAMP phosphodiesterase, which seems to be functionally closely coupled with the hormone-sensitive lipase-regulatory system influencing primarily a pool of cAMP utilized by the relevant protein kinase.

3T3-L1 adipocytes contain both soluble and particulate cAMP phosphodiesterases which can be distinguished by several criteria. Particulate phosphodiesterase activity of 3T3-L1 adipocytes, but not undifferentiated fibroblasts, was selectively increased by incubation of cells with insulin or lipolytic hormones. Particulate cAMP phosphodiesterase activity from 3T3-L1 adipocytes was very sensitive to inhibition by cilostamide in an apparently competitive fashion. Particulate activity from undifferentiated 3T3-L1 fibroblasts or supernatant activity from either type of cell was much less sensitive to cilostamide. On the other hand, supernatant cAMP phosphodiesterase activity from both undifferentiated fibroblasts and 3T3-L1 adipocytes was very sensitive to inhibition by Ro-20-1724 in an apparently competitive fashion. Ro-20-1724 was not an effective inhibitor of particulate activity from either type of cell. In fractions from 3T3-L1 adipocytes, isobutylmethylxanthine (IBMX) effectively inhibited both supernatant and particulate cAMP phosphodiesterase activities. In addition, however, IBMX was relatively more specific in inhibiting supernatant calmodulin-activated cGMP phosphodiesterase activity than supernatant calmodulin-independent or particulate cGMP phosphodiesterase activities. In intact 3T3-L1 adipocytes, cilostamide enhanced lipolysis in the absence or presence of isoproterenol and had no effect on cAMP content in the presence of low concentrations of isoproterenol. Ro-20-1724 increased lipolysis to a lesser extent than cilostamide and did not enhance isoproterenol-stimulated lipolysis, but did increase isoproterenol-stimulated accumulation of cAMP to a greater extent than cilostamide. Like cilostamide, Ro-20-1724 did not enhance accumulation of cAMP in the absence of isoproterenol. IBMX enhanced lipolysis and cAMP accumulation with or without isoproterenol. Taken together, these results support the idea that although particulate and soluble low Km phosphodiesterases influence cAMP content, the particulate enzyme may be more important in the metabolism of cAMP involved in the regulation of lipolysis. Since combinations of Ro-20-1724 and cilostamide were not as effective as IBMX in increasing cAMP content, perhaps the calmodulin-dependent phosphodiesterase, which is selectively inhibited by IBMX, is also involved in the regulation of total cell cAMP content.

No abstract

By the WestJEM article submission agreement, all authors are required to disclose all affiliations, funding sources and financial or management relationships that could be perceived as potential sources of bias. No author has professional or financial relationships with any companies that are relevant to this study. There are no conflicts of interest or sources of funding to declare.

We have used cultured 3T3-L1 adipocytes to assess direct effects of T3 on beta-adrenergic-mediated regulation of lipolysis and adenylate cyclase and phosphodiesterase activities. Differentiated 3T3-L1 adipocytes were maintained under four conditions: in the presence of medium containing serum from a hypothyroid calf (hypothyroid medium), hypothyroid medium supplemented with T3 (T3-supplemented hypothyroid medium), medium with serum from a normal calf (control medium), and control medium supplemented with excess T3 (hyperthyroid medium). Compared to the two control groups, i.e. cells maintained in control medium or T3-supplemented hypothyroid medium, cells maintained in hypothyroid medium exhibited lower basal rates of lipolysis and lower sensitivity to isoproterenol. Hyperthyroid cells exhibited higher basal rates of lipolysis and higher sensitivity to isoproterenol. In the presence of maximally effective concentrations of isoproterenol, rates of lipolysis were similar in the four groups. Similarly, basal cAMP content and cAMP accumulation in the presence of isoproterenol were reduced in hypothyroid and increased in hyperthyroid adipocytes compared to those adipocytes maintained in control or T3-supplemented hypothyroid medium. Basal adenylate cyclase activity was similar in the four groups. Sensitivity to isoproterenol and maximal isoproterenol-stimulated cyclase activity were diminished in membrane preparations from hypothyroid adipocytes and increased in preparations from hyperthyroid adipocytes. Cyclase activity in the presence of NaF, however, was similar in preparations from cells maintained in hypothyroid, T3-supplemental hypothyroid, or control medium. NaF-stimulated activity was increased in preparations from hyperthyroid adipocytes. Thyroid status did not affect beta-receptor number of affinity for iodohydroxybenzylpindolol. Compared to control cells or cells maintained in T3-supplemented hypothyroid medium, both soluble and particulate cAMP phosphodiesterase activities were increased in hypothyroid cells and decreased in hyperthyroid cells. These results indicated that in 3T3-L1 adipocytes, some of the effects of thyroid hormone on cAMP content and lipolysis can be explained by alterations in both production and degradation of cAMP.