Topiramate treatment causes skeletal muscle insulin sensitization and increased Acrp30 secretion in high-fat-fed male Wistar rats

Jason J. Wilkes, M. T. Audrey Nguyen, Gautam K. Bandyopadhyay, Elizabeth Nelson, and Jerrold M. Olefsky

Division of Endocrinology and Metabolism, Department of Medicine, University of California, San Diego; and The Whittier Diabetes Institute, La Jolla, California

Submitted 18 April 2005 ; accepted in final form 18 July 2005


    ABSTRACT
 TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
We show that Topiramate (TPM) treatment normalizes whole body insulin sensitivity in high-fat diet (HFD)-fed male Wistar rats. Thus drug treatment markedly lowered glucose and insulin levels during glucose tolerance tests and caused increased insulin sensitization in adipose and muscle tissues as assessed by euglycemic clamp studies. The insulin-stimulated glucose disposal rate increased twofold (indicating enhanced muscle insulin sensitivity), and suppression of circulating FFAs increased by 200 to 300%, consistent with increased adipose tissue insulin sensitivity. There were no effects of TPM on hepatic insulin sensitivity in these TPM-treated HFD-fed rats. In addition, TPM administration resulted in a three- to fourfold increase in circulating levels of total and high-molecular-weight (HMW) adiponectin (Acrp30). Western blot analysis revealed normal AMPK (Thr172) phosphorylation in liver with a twofold increased phospho-AMPK in skeletal muscle in TPM-treated rats. In conclusion, 1) TPM treatment prevents overall insulin resistance in HFD male Wistar rats; 2) drug treatment improved insulin sensitivity in skeletal muscle and adipose tissue associated with enhanced AMPK phosphorylation; and 3) the tissue "specific" effects are associated with increased serum levels of adiponectin, particularly the HMW component.

high fat; insulin sensitization; adenosine monophosphate-activated protein kinase; adiponectin


INSULIN RESISTANCE is a characteristic feature of type 2 diabetes and is the defining abnormality of syndrome X, or the metabolic syndrome. Given the epidemic proportion of these disorders, development of new insulin-sensitizing therapeutics could be of great importance. Topiramate is a clinically approved antiseizure compound, and clinical experience as well as limited animal studies have suggested that this agent may lead to decreased insulin resistance and modest weight loss. In earlier studies (23), we showed that treatment of female Zucker fatty (ZDF) rats with Topiramate led to improved insulin sensitivity, independently of weight loss, and that the primary drug effects were exerted on adipose tissue. Thus overall in vivo insulin sensitivity was improved, but when tissues were excised and studied ex vivo, adipose tissue insulin sensitization was maintained, but skeletal muscle insulin action was no longer enhanced. Consistent with this, direct in vitro studies showed that Topiramate caused insulin sensitization when added to fat tissue, but not muscle.

Adiponectin (Acrp30) is synthesized and secreted exclusively by adipocytes (16), and many studies have shown that high levels of adiponectin are associated with insulin sensitivity whereas low levels are found in insulin resistance (8, 9, 17). Furthermore, direct experiments have demonstrated that adiponectin can mediate insulin-sensitizing effects, most likely by activating AMP-activated protein kinase (AMPK).

Adiponectin circulates in a low-molecular-weight (LMW) hexameric form and a high-molecular-weight (HMW) form consisting of trimer hexamers, and it is the HMW form of circulating adiponectin that may contain most of the in vivo insulin-sensitizing activity (10, 11, 21). This adipokine appears to exert its biological effects by binding to its recently identified cell surface R1 and R2 receptors (24) followed by activating of AMPK (25). Recently, we have shown that adenoviral mediated ectopic transgenic expression of adiponectin improves whole body insulin sensitivity in Wistar rats, completely protects these animals from high-fat diet (HFD)-induced insulin resistance, and leads to activation of AMPK activity in skeletal muscle, but not liver.

On this basis, we speculated that Topiramate-induced insulin sensitization may be mediated, at least in part, through adipocyte-derived adiponectin. The current studies in HFD Wistar rats treated with Topiramate provide strong support for this hypothesis.


    MATERIALS AND METHODS
 TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
Materials. Anti-AMPK{alpha} and phospho-AMPK (Thr172) antibodies were purchased from Cell Signaling Technology (Beverly, MA). PVDF membrane was purchased from Millipore (Bedford, MA). HFD was purchased from DyEts (Bethlehem, PA). General reagents were purchased from Sigma Chemical (St. Louis, MO). Johnson & Johnson (Raritan, NJ) kindly provided us with Topiramate (TPM).

General use of animals. Male Wistar rats (Charles River, Wilmington, MA) weighing between 350 and 375 g were used. To induce obesity, we divided rats into two groups and provided one with chow and the other an HFD. Animals on HFD were given a diet that contained saturated fat as the primary source of calories (55% hydrogenated coconut oil, 15% casein, 15% sucrose, 4% soybean oil), whereas rats in the low-fat group were given normal rodent chow with <5% dietary fat. HFD rats were permitted to consume their diet freely for 5–6 wk. Thereafter, pair feeding was implemented to control for the food restriction effects of TPM. Thus rats on HFD were matched by body weight and allotted to pair-fed HFD + placebo, regular fed HFD + TPM or untreated regular feeding HFD group. Pair-fed rats consumed equal portions of food as TPM-treated rats daily. Single daily doses of TPM (100 mg/kg) were given by oral gavage to TPM-treated rats. Drug-treated (HFD regular fed + TPM) and placebo treated (HFD pair fed + placebo) and normal chow-fed animals all underwent glucose tolerance testing on day 16 to generate data on general aspects of insulin sensitivity (i.e., glucose and insulin levels) of experimental groups. More detailed assessments of muscle and liver insulin sensitivity [i.e., glucose disposal rate (GDR), hepatic glucose production (HGP)] were made in fully recovered catheterized rats that were pair fed and untreated or regular fed and treated with and without TPM.

Glucose tolerance tests. Glucose tolerance tests (GTT) were performed in overnight-fasted rats on HFD and treated with or without Topiramate, as well as in untreated chow-fed animals. First, unchallenged blood draws were performed for basal glucose and insulin measurements. This was achieved by nipping tails and collecting blood from tail veins (~100 µl per rat) in heparinized capillary tubes. Glucose concentrations were determined immediately using a portable blood glucose analyzer (Hemocue, Mission Viejo, CA). Plasma was spun out of remaining samples and stored at –80°C. Plasma insulin was determined at a later time. After basal sampling, glucose loads were delivered (3 g/kg body wt ip) to hand-held conscious animals. Thereafter, time points for glucose measurements were made at 60, 120, and 180 min and insulin at 60 and 180 min.

Surgery and clamp procedure. Rats were inserted with cannulae on day 16 of the protocol in the same manner as we have described previously (22). Euglycemic hyperinsulinemic clamp studies were performed 7 to 9 days after cannulation in fasted rats, as before, but with one minor technical modification. Instead of arterial blood sampling, blood was collected via the tail vein to be consistent with the sampling technique used in our GTT procedure. Basal blood samples were drawn at –60 and 0 min. A priming dose (5 µCi) of D-[3-3H]glucose tracer (New England Nuclear, Boston, MA) was administered by bolus injection. Then (at –60 min) tracer was administered by constant infusion (0.167 µCi/min) for 1 h to further assist in the equilibration of tritium with the glucose pool. At 0 min, a solution of cold glucose (50% dextrose; Abbott Labs, Chicago, IL) was infused at a variable rate along with the infusion of tritium plus insulin (25 mU·kg–1·min–1 Novlin R; Novo Nordisk, Copenhagen, Denmark) which was infused at a fixed rate (16.7 µl/min). Variable and fixed infusions of glucose and tritium plus insulin solutions were started simultaneously and maintained throughout the duration of the clamp. Glucose pumps were adjusted as needed to correct for changes in blood glucose levels during insulin stimulation. This was achieved by manually adjusting pump rates every 10 min until euglycemia was reached. Blood samples were taken at the completion of the 2-h clamp for determination of glucose, insulin, free fatty acids (FFA), adiponectin, and lactate. All blood samples were immediately centrifuged, and plasma was stored at –80°C. A terminal dose of Nembutal (100 mg/kg iv) was administered after clamping to dissect liver, as well as extensor digitorum longus (EDL) muscle, from euthanized rats.

Analysis of plasma hormones and metabolites. Plasma insulin and plasma adiponectin were measured via double-antibody radioimmunoassay techniques specific for insulin and adiponectin, respectively (LINCO Research, St. Charles, MO). Plasma FFA was measured enzymatically using a commercially available kit (NEFA C; Wako Chemicals USA, Richmond, VA). Plasma lactate was determined with a YSI 1500 SPORT plasma analyzer fitted with YSI 2329 membranes suitable for L-lactate analysis (Yellow Springs Instrument, Yellow Springs, OH).

Analysis of the oligomeric distribution of serum adiponectin. Plasma samples (2 µl) were fractionated by PAGE on a 4–15% gel under nonreducing conditions, transferred onto PVDF membrane (Immobilon-P, Millipore), and blotted with a rat adiponectin antibody (Affinity BioReagents, Golden, CO). Membranes were then incubated with horseradish peroxidase-conjugated secondary antibody before chemiluminescence detection (Pierce, Rockford, IL). Band intensities for LMW and HMW adiponectin were quantified by densitometry using a Macintosh computer connected to an ARCUS scanner by way of NIH Image 1.6 software.

AMPK measurement in muscle and liver tissues. EDL and liver samples underwent homogenization to generate tissue lysates for phospho-AMPK Western blotting, as described in a previous paper (22). Western blotting was performed using acrylamide-based gels. Visualization of Western blots was made possible by an enhanced chemiluminescence system (Pierce). Bands were quantified using a Macintosh computer connected to an ARCUS scanner by way of NIH Image 1.6 software.

Muscle triglyceride analysis. Lipids were extracted from muscle by a method adapted from Frayn and Maycock (7). Total intramuscular triglyceride (IMTG) content was determined in lipid extracts by use of a kit from Thermo DMA (Arlington, TX).

Calculations and statistical analysis. HGP and GDR were calculated using Steele's equation (18). All data were analyzed using two-way analysis of variance and t-tests as appropriate. Data calculation and statistical analysis were performed using StatView (Abacus Concepts, Berkley, CA). All data are reported as means ± SE.


    RESULTS
 TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
GTTs. TPM has a transient effect to suppress food intake in rats, resulting in transient weight loss (23). Therefore, to study rats at an equal body weight, we first induced obesity (i.e., before dosing) by HFD and then matched food intakes of TPM- and placebo-treated groups by pair feeding. Fig. 1A shows that the HFD caused rats to gain weight and that pair feeding effectively normalized body weights of placebo- and TPM-treated rats. Table 1 shows that rats treated with TPM and those that were pair fed consumed equal amounts of food over the final 2–3 days of treatment. As seen in Fig. 1B, both groups of rats on HFD (HFD + placebo and HFD + TPM) showed identical body weights, and both were significantly (P < 0.05) heavier (15–20%) than regular chow-fed rats. Thus we performed GTTs in moderately obese rats as well as in normal chow-fed animals, and the results, as shown in Fig. 1, indicate that TPM treatment prevented the HFD-induced glucose intolerance (Fig. 1C) and also lowered basal and postprandial insulin levels (Fig. 1D).



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Fig. 1. Glucose tolerance is normalized in Topiramate (TPM)-treated high-fat-diet (HFD)-fed obese Wistar rats. A: HFD feeding increased body weights of Wistar rats. Pair feeding normalized body weights of TPM- and placebo-treated rats. All experimental groups of rats underwent glucose tolerance testing (3 g/kg ip) on day 16. B: body weights of HFD-fed rats were increased by day 16; *P < 0.05 vs. chow-fed rats. C and D: consumption of HFD impaired glucose tolerance, and TPM treatment normalized glucose-challenged plasma glucose (C) and plasma insulin (D) concentrations in basal and postprandial states; ¶P < 0.05 vs. placebo-treated controls. Bars represent means ± SE (n = 8).

 

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Table 1. Food record of HFD rats on TPM treatment or placebo

 
Euglycemic clamp studies. To confirm the results of the GTTs showing that TPM treatments improve whole body glucose homeostasis in HFD Wistar rats, we subjected all three groups to euglycemic hyperinsulinemic clamp studies. Table 2 shows some general characteristics of Wistar rats carrying indwelling catheters and receiving treatments of TPM or placebo in the postsurgical state. As seen in Table 2, chronically catheterized Wistars on HFD and undergoing treatments with or without TPM were equal in body size at the time of hyperinsulinemic euglycemic testing and significantly heavier than normal rats (P < 0.05). All groups of cannulated rats on HFD displayed modest but significant increases (P < 0.05) in basal blood glucose (+1.5 -2 mM) and basal plasma insulin (+1–1.5 ng/ml) concentrations compared with normal chow-fed controls. Clamped glucose and clamped lactate levels were identical among all groups of rats (HFD, HFD + TPM, chow). Basal plasma FFAs were equal among HFD, HFD + TPM, and chow groups. Plasma FFAs are normally suppressed during hyperinsulinemic euglycemic clamp studies, and an overall suppression of serum FFAs occurred in all three groups of rats (P < 0.05 for glucose clamped vs. basal). TPM-treated rats showed a greater suppression of serum FFAs than HFD controls (P < 0.05) and demonstrated significantly lower serum FFA levels during glucose clamps compared with HFD controls (P < 0.05).


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Table 2. Metabolic characteristics of catheterized male Wistar rats on a regular chow diet or HFD and receiving a 100 mg/kg daily dose of TPM (HFD + TPM) or placebo (HFD + PL) for 7–9 days

 
Figure 2 shows the glucose infusion rates required to maintain steady-state blood glucose concentrations in the clamped rats from the experimental groups. As seen in Fig. 2, HFD led to a 25% decrease in the glucose infusion rate (GINF), whereas the GINF was increased by 25–40% (P < 0.05) in HFD animals on TPM compared with HFD rats treated with placebo. The GINF in TPM-treated rats (40–45 mg·kg–1·min–1) was similar to the GINF of chow-fed rats (40–45 mg·kg–1·min–1) and significantly higher (P < 0.05) than the GINF in HFD control rats (30–35 mg·kg–1·min–1).



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Fig. 2. Reversal of diet-induced whole body insulin resistance as determined by glucose infusion rate (GINF). HFD decreased GINF measured in mg·kg–1·min–1; *P < 0.05 vs. chow. TPM-treated rats on HFD demonstrated reversal of whole body insulin resistance, as shown by reversal of GINF. The GINF of rats on TPM treatment was less than those on placebo (¶P < 0.05) and equal to those on chow. Bars represent means ± SE (n = 6).

 
Next, we assessed the effects of TPM treatment on muscle and liver insulin action by calculating [3H]glucose tracer-derived GDR and HGP. As seen in Fig. 3A, HFD led to insulin resistance resulting in the expected decrease in GDR, whereas TPM treatment led to an increase in GDR in the HFD rats. Because the majority of GDR occurs in skeletal muscle, these results indicate a marked enhancement of in vivo muscle glucose disposal in TPM rats compared with the HFD group and the chow-fed rats. With respect to liver, there was no improvement in hepatic insulin sensitivity resulting from TPM treatment (Fig. 3B). Figure 3B shows that hepatic insulin resistance developed in both treated and untreated HFD animals, as shown by a decreased effect of insulin to suppress HGP.



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Fig. 3. TPM treatment ameliorates effect of diet in muscle without correcting insulin resistance in liver. Glucose disposal rates [GDR and insulin-stimulated (IS)-GDR], are shown in A and C, respectively. Hepatic glucose production (HGP) is shown in B. Metabolic effects of insulin were determined by hyperinsulinemic clamp in combination with [3H]glucose infusion and analysis of tracer in dehydrated plasma. A: TPM treatment increased GDR in HFD-fed rats; ¶¶P < 0.01 vs. HFD-fed controls; *P < 0.05 vs. chow-fed. B: HGP suppression by insulin was impaired by HFD, and insulin-stimulated HGP was impaired in HFD + TPM-treated rats; *P < 0.05 vs. chow. C: net effect of insulin on in vivo glucose uptake (IS-GDR). IS-GDR was reduced in HFD rats; *P < 0.05 vs. chow. Also, HFD + TPM-treated rats had increased IS-GDR; ¶P < 0.05 vs. HFD-fed controls, and IS-GDR in TPM animals was similar to that in normal animals. Single filled bars represent means ± SE of GDR and IS-GDR. Double bars are for basal (filled) and insulin-stimulated (open) levels of HGP (n = 6).

 
To more accurately quantitate the effect of TPM treatment on muscle insulin sensitivity, we calculated insulin-stimulated GDR (IS-GDR). IS-GDR is typically used to illustrate the net effect of insulin on in vivo muscle glucose uptake (IS-GDR = GDR – basal HGP). As shown in Fig. 3C, the HFD-induced decrease in IS-GDR was completely prevented by TPM treatment.

Circulating adiponectin levels. Adiponectin is an adipocyte-secreted peptide with an ability to augment insulin sensitivity, and previous studies have shown that circulating adiponectin levels are low in insulin-resistant states, such as HFD, and are upregulated when insulin sensitivity is improved by treatment with thiazolidinediones (1). To assess the effects of TPM treatment on adiponectin secretion, total adiponectin was measured in the experimental groups. As seen in Fig. 4A, HFD led to a decrease in total adiponectin levels, whereas TPM treatment led to a marked increase.



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Fig. 4. Circulating levels of adiponectin (Acrp30) are increased in TPM-treated rats. Rats were subjected to a 2-h hyperinsulinemic clamp as described in MATERIALS AND METHODS. Plasma was collected afterward, stored at –80°C, and, upon completion of all studies, analyzed for total Acrp30 protein by RIA. TPM-treated rats displayed significantly increased plasma levels of total Acrp30 compared with placebo-treated (¶P < 0.01) and chow-fed controls. Bars represent means ± SE (n = 6). C, chow; HF, HFD; HMW and LMW, high- and low-molecular-weight Acrp30, respectively. *P < 0.05.

 
AMPK phosphorylation and IMTG content. Pharmacological studies (2, 19) have shown that EDL skeletal muscles and primary hepatocytes respond directly to adiponectin. Therefore, EDL and liver sections were harvested from clamped animals and prepared, as described in MATERIALS AND METHODS, for quantification of total AMPK{alpha} and phospho-AMPK levels.

Figure 5 shows that AMPK protein levels and AMPK phosphorylation in liver tissue were the same among all study groups. In Fig. 6A, it can be seen that the level of phospho-AMPK in EDL skeletal muscle was decreased by HFD (P < 0.05) and that this was reversed by TPM treatment, resulting in an increase to a level above that of normal animals (P < 0.05). The HFD also led to an approximately twofold increase in IMTG, which was completely prevented when the HFD-fed animals were treated with TPM (Fig. 6B). This is fully consistent with the TPM-enhanced increase in muscle AMPK activity and insulin sensitivity.



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Fig. 5. AMP-activated protein kinase (AMPK) phosphorylation (P-AMPK) in liver in TPM-treated rats. TPM-treated rats displayed normal AMPK phosphorylation levels compared with HFD-fed and regular (chow) controls.

 


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Fig. 6. A: P-AMPK is upregulated in muscle in TPM-treated rats. EDL muscles harvested at the end of the euglycemic clamp studies were analyzed for AMPK{alpha} and P-AMPK (Thr172). TPM-treated rats displayed significantly increased in vivo levels of AMPK phosphorylation; ¶¶P < 0.01 vs. HFD-fed controls; *P < 0.05 vs. regular fed controls. B: intramuscular triglyceride (IMTG) levels were increased by HFD (*P < 0.05 vs. chow) and decreased by TPM treatment (¶P < 0.05 vs. HFD alone). Bars represent means ± SE (n = 5–6).

 

    DISCUSSION
 TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
Insulin resistance is a central feature of type 2 diabetes as well as a number of other common human insulin-resistant states; therefore, identification of a new insulin-sensitizing therapeutic would be of great importance. Topiramate is a currently marketed antiepileptic agent that has been shown to cause weight loss and improve insulin sensitivity in both humans (4, 13) and animals (12, 14). Our own studies have shown that Topiramate treatment in insulin-resistant female ZDF rats can lead to amelioration of insulin resistance independently of weight loss. Although overall in vivo insulin sensitivity was improved in these animals, when tissues were studied ex vivo adipose tissue retained its insulin-sensitive state, whereas the effects in skeletal muscle were lost after removal from the in vivo environment. This led us to conclude that Topiramate leads to insulin sensitization, exerting its effects predominantly in adipose tissue.

Adiponectin is an adipocyte-secreted factor that can cause insulin sensitization in a variety of conditions (6, 26). This adipokine circulates in HMW and LMW components, and recent evidence suggests that it is the HMW fraction of adiponectin that is primarily responsible for the insulin sensitizing effects, presumably by binding to its G protein-coupled receptor-like cell surface receptor and activating AMPK in target tissues. We have recently found (15) that ectopic expression of adiponectin, using adenoviral mediated gene transfer, leads to increased insulin sensitivity and complete protection from the effects of HFD to cause insulin resistance. Taken together, these results led us to speculate that Topiramate may exert its insulin-sensitizing effects through an adiponectin-related mechanism.

In the current study, we treated HFD-fed male Wistar rats with Topiramate or placebo, followed by measurement of glucose tolerance, insulin sensitivity, adiponectin levels, and tissue AMPK activity. The major findings from this study are that Topiramate treatment prevents HFD-induced glucose intolerance and hyperinsulinemia as measured during GTTs. Hyperinsulinemic euglycemic clamp studies showed that HFD led to insulin resistance, as shown by a 40% decrease in the IS-GDR. Because the great majority of overall GDR is into skeletal muscle, this result shows HFD-induced skeletal muscle insulin resistance. This HFD-induced decrease in IS-GDR was completely prevented by Topiramate, and, in fact, IS-GDR values in HFD + TPM animals were somewhat higher than in chow-fed controls. To the extent that suppression of FFA levels during the clamp procedure reflect insulin's antilypolytic effects, the HFD impaired, whereas Topiramate treatment enhanced, adipose tissue insulin sensitivity. Interestingly, HFD also led to hepatic insulin resistance as manifested by an impaired ability of insulin to suppress HGP, and this was not affected by Topiramate treatment.

Our experiments also reveal potential mechanisms to explain these Topiramate effects. Thus we measured adiponectin in all of the experimental groups. As seen in Fig. 4, HFD causes a decrease and Topiramate treatment leads to an increase in total circulating adiponectin levels compared with chow-fed controls.

Adiponectin is known to stimulate AMPK activity, and this could result in the enhanced fat oxidation (19, 25) and all of the other insulin-sensitizing effects of adiponectin that have been described. We measured AMPK activity in muscle and liver tissue from these animals and found enhanced AMPK phosphorylation in muscle, but not in liver, in the HFD + TPM animals, consistent with the in vivo effects of Topiramate on IS-GDR but not HGP. Additionally, plasma FFAs were found to be lower after insulin stimulation in glucose-clamped HFD + TPM animals compared with controls, which could have also affected muscle insulin sensitivity. Interestingly, the HFD-induced accumulation of IMTG was also prevented by Topiramate treatment.

Topiramate has been reported to cause decreased food intake and weight loss, which are largely transient in rodents. Indeed, there was a modest difference in body weights of Topiramate-treated and untreated non-pair-fed HFD animals. However, this was not a factor in our studies, since all comparisons were made between HFD + TPM-treated animals and pair-fed HFD + placebo groups, and food intake and body weights were identical between these two groups (Table 1 and Fig. 1A). Indeed, over the last 24 h of the study, food intake was only ~1 g/day less in the Topiramate-treated and pair-fed placebo groups compared with the ad libitum-fed HFD animals.

The literature contains mixed results as to the tissue site of adiponectin's insulin-sensitizing effects. Some studies have shown effects predominantly in liver (2, 5) while others have clearly shown direct adiponectin actions on skeletal muscle (25, 3). Studies, for example, with 5-h-fasted mice given recombinant adiponectin (20 ng·g body wt–1·min–1) to acutely elevate plasma adiponectin concentrations showed that adiponectin improves hepatic insulin sensitivity without improving peripheral insulin sensitivity. These hepatic effects were associated with reductions in glucose flux through the glucose-6-phosphatase metabolic pathway, with no effect on glycolysis or glycogen synthesis. Indeed, others have also reported on adiponectin's liver selective actions. Berg et al. (2) showed that the full-length adiponectin protein has glucose-lowering effects in ob/ob and nonobese diabetic mice, and also demonstrated direct effects of full-length adiponectin on isolated primary hepatocytes. In contrast, globular adiponectin has direct effects in muscle to activate AMPK, stimulate glucose uptake and FFA oxidation (25), and cause GLUT4 translocation in L6 cells (3). In our own earlier studies (15), using adenovirus gene transfer to achieve ectopic transgenic expression of adiponectin in Wistar rats on HFD, we observed that the predominant insulin-sensitizing effects of the expressed adiponectin were exerted in skeletal muscle, as demonstrated by increased in vivo IS-GDR and enhanced muscle AMPK activity. This is similar to the current studies, where we find that Topiramate treatment leads to enhanced muscle, but not liver, insulin sensitivity. Although the reason for the differences between various animal studies is unknown, it is possible that they may be partially related to differences in adiponectin receptor expression. For example, Yamauchi et al. (24) identified two adiponectin receptors, Adipo-R1 and Adipo-R2. In mice, Adipo-R1 is expressed predominantly in muscle, with Adipo-R2 highly expressed in liver, and exogenous adiponectin exerts potent effects to enhance hepatic insulin sensitivity in mice. In rats, both Adipo-R1 and -R2 are expressed more highly in muscle, possibly explaining adiponectin-induced muscle insulin-sensitizing effects in rats. Interesting, the pattern of adiponectin receptor expression that we observed in rats is comparable to the pattern reported in humans, raising the possibility that Topiramate may work in man similarly to rats.

In conclusion, these studies show that Topiramate treatment prevents HFD-induced insulin resistance in male Wistar rats. This is accompanied by a drug-induced increase in adipose tissue-derived circulating HMW adiponectin, with increased skeletal muscle AMPK activity. Taken together, these results suggest that Topiramate treatment can enhance muscle insulin sensitivity, and this may reflect a direct effect on adipocytes to increase adiponectin secretion.


    GRANTS
 TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
This work was supported by research grants from the National Institute of Diabetes and Digestive and Kidney Diseases (DK-33651) and Johnson and Johnson Pharmaceutical Research & Development.


    ACKNOWLEDGMENTS
 
Betsy Hansen assisted in the preparation of the manuscript.


    FOOTNOTES
 

Address for reprint requests and other correspondence: J. Wilkes, Dept. of Medicine (0673), UCSD, 9500 Gilman Dr., La Jolla, CA 92093 (e-mail: jwilkes{at}gnf.org)

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.


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 DISCUSSION
 GRANTS
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