The adipose hormone leptin potently influences physical activity. Leptin can decrease locomotion and running, yet the mechanisms involved and the influence of leptin on the rewarding effects of running (“runner’s high”) are unknown. Leptin receptor (LepR) signaling involves activation of signal transducer and activator of transcription-3 (STAT3), including in dopamine neurons of the ventral tegmental area (VTA) that are essential for reward-relevant behavior. We found that mice lacking STAT3 in dopamine neurons exhibit greater voluntary running, an effect reversed by viral-mediated STAT3 restoration. STAT3 deletion increased the rewarding effects of running whereas intra-VTA leptin blocked it in a STAT3-dependent manner. Finally, STAT3 loss-of-function reduced mesolimbic dopamine overflow and function. Findings suggest that leptin influences the motivational effects of running via LepR-STAT3 modulation of dopamine tone. Falling leptin is hypothesized to increase stamina and the rewarding effects of running as an adaptive means to enhance the pursuit and procurement of food.

Leptin modulates multiple components of brain reward circuitry (), and LepR signaling in dopamine (DA) neurons of the ventral tegmental area (VTA) is well-documented (). LepR knockdown in the VTA increases food intake and locomotor activity () and food-motivated behavior (). LepR signaling includes the activation of signal transducer and activator of transcription 3 (STAT3) (), which is observed in DA and GABA neurons of the VTA (). However, leptin-induced pSTAT3 is found in only a subset of LepR-positive neurons (), suggesting that LepR-STAT3 signaling may have unique neurobehavioral actions. Given the implication of ERK signaling in the anorectic actions of VTA leptin (), here we tested the hypothesis that STAT3 in DA neurons is involved in the control of physical activity, including voluntary running and the rewarding effects of running.

Physical activity is strongly influenced by metabolic state. The adiposity hormone leptin controls feeding and energy expenditure, and a fall in leptin is a major component of the physiological response to fasting (). Leptin increases locomotor activity and voluntary running during fed states in leptin-deficient humans (), ob/ob mice (), wild-type mice (), and rats (), or has no effect on running in fed wild-type mice (). In contrast, leptin inhibits locomotor activity and voluntary wheel running when food is limited in ob/ob and lean mice () and rats (). Physical activity escalates with food restriction, and leptin concentrations are inversely related to running in humans (), rats (), wild-type mice (), and in mice bred for high running capacity (). Furthermore, leptin levels inversely correlate with marathon run-time independent of BMI () and with run speed and duration in high-running mice (), suggesting that leptin impacts the motivational and rewarding effects of running. Indeed, endurance running is rewarding in humans (“runners high”) () and rodents (), yet little is known about the neural mechanisms involved.

All behavior entails energy expenditure in the form of physical activity. Activity that increases the likelihood that a physiological need will be met is referred to as appetitive, and appetitive behaviors are crucial to the fulfillment of energy demands. Food-directed appetitive behaviors such as foraging and hunting benefit from increased activity and stamina. The capacity for endurance running in mammals is considered to have evolved as a means to increase the payoff of food-directed behaviors targeting distant or shifting food sources ().

The NAc is strongly innervated by VTA DA neurons, and leptin deficiency reduces NAc DA overflow and TH, the rate-limiting enzyme for DA biosynthesis (). We used fast-scan cyclic voltammetry ( Figure 4 A), a real-time electrochemical method, to measure axonal DA overflow in the NAc core sub-compartment that has been linked to the control of wheel running (). Evoked DA overflow was significantly diminished in STAT3mice ( Figures 4 B and 4C). In addition, TH and D1 receptor expression were decreased in the NAc of STAT3mice ( Figure 4 D), similar to findings in mice bred for high-running capacity (). We next assessed the locomotor-stimulating effects of a DA D1 receptor agonist and locomotor sensitization in response to amphetamine (AMPH). STAT3mice displayed decreased locomotion in response to D1 receptor agonism ( Figures 4 E and 4F) and failed to sensitize to AMPH ( Figures 4 G and 4H). Reduced DA overflow in the NAc is reported in conditions of increased food reward, including diet-induced obesity () and leptin deficiency (). Similarly, blunted responses to AMPH and compromised striatal DA signaling are tied to enhanced reward seeking in humans (). As a final mesure, we measured the expression of several opioid and eCB markers in the VTA and NAc. STAT3mice exhibited a near-significant increase in delta opioid receptor (DOR) mRNA levels in the NAc ( Figure 4 I). Higher DOR levels could amplify opioid signaling to increase running reward. Consistent with increased opioid tone during the runner’s high, elevated dynorphin mRNA levels in the NAc core have been observed in a high-running rat strain (), a trend observed for pre-prodynorphin mRNA in STAT3mice ( Figure 4 I). In contrast, pre-prodynorphin mRNA was attenuated in the VTA of STAT3mice ( Figure 4 J). While eCBs themselves were not measured, no changes were found in any eCB-related molecules.

(B) Current-time plot showing subsecond stimulation-evoked DA release and reuptake. Oxidation current peaks for DA were obtained at potentials of −300 to −500 mV (versus Ag/AgCl) corresponding to 3.5–4.5 ms in the voltage waveform.

Endurance running or exercise can promote a lasting sense of well-being that includes feelings of euphoria and pleasantness commonly referred to as the runner’s high (). Running also has rewarding properties in rodents: rats will work to gain access to a running wheel () and spend more time in a place associated with the aftereffects of wheel running (). Addictive and compulsive-like behavior directed at high levels of physical activity are reported in humans () and rodents (). Running engages opioid (), DA () and endocannabinoid (eCB) systems (). Using a conditioned place preference (CPP) task ( Figure 3 A), we found that the rewarding effect of running is increased in KO mice: STAT3mice spend more time in the paired side of the chamber associated with running ( Figures 3 B and 3C). Next, a separate cohort of mice received either intra-VTA saline or leptin at a dose shown to inhibit feeding ( Figure 2 A) prior to CPP testing. STAT3mice treated with vehicle again displayed increased running reward ( Figure 3 D). VTA leptin completely suppressed the rewarding effects of running in controls yet was unable to block running reward in STAT3mice ( Figure 3 E).

We next ascertained if STAT3 signaling in DA neurons mediates the anorectic effects of central leptin. Decreases in food intake following intra-VTA leptin were not different between control and STAT3mice ( Figure 2 A). Likewise, reductions in food intake following ICV injection of leptin were similar ( Figure 2 B), suggesting that LepR-STAT3 signaling in DA neurons has a minimal impact on the anorectic actions of leptin. We then employed an operant task to determine the role of STAT3 signaling in DA neurons in food-motivated behavior. STAT3mice exhibited significant response impairments as evidenced by reduced lever presses ( Figure 2 C) and increased percentage of incorrect responses ( Figure 2 D). Elevated incorrect responses could reflect a compulsive phenotype. To address this possibility, we performed a conditioned suppression test to evaluate sucrose intake in the context of aversive conditioning. Sucrose intake was similarly suppressed across groups implying that STAT3mice are not compulsive eaters ( Figure 2 E). As a final approach, we employed a hedonic feeding test whereby sated mice overeat palatable food (). Caloric intake increased to a similar extent in both groups when exposed to the sweetened, high-fat food (dessert) ( Figure 2 F).

STAT3mice weighed significantly less than controls as of 19 weeks of age ( Figure 1 A), an effect associated with less fat mass ( Figure 1 B). Normalized caloric intake was comparable between genotypes ( Figure 1 C), but feed efficiency was decreased in STAT3mice ( Figure 1 D) to suggest elevated energy expenditure. Heat dissipation and respiratory exchange were unchanged ( Figures 1 E and 1F); thus, we next explored if heightened physical activity contributes to greater energy expenditure. Horizontal movement ( Figure 1 G) and distance traveled in the dark cycle (inset) were elevated in STAT3mice. We next assessed voluntary wheel running to find that it was substantially increased in STAT3mice ( Figure 1 H), which ran ∼11 km/day as compared to ∼6 km/day of controls ( Figure 1 I). Running was similar between DATmice and littermates controls, suggesting that increased running of STAT3mice is not attributable to Cre transgene alone ( Figure S2 ). We next rescued STAT3 via VTA microinjections of an adeno-associated virus (“AAV-DIO-STAT3”; Figure 1 J). The STAT3 transgene was expressed only in DA neurons ( Figures S3 A and S3B) and increased STAT3 expression ( Figure S3 C). AAV-DIO-STAT3 normalized voluntary running of STAT3to that of controls ( Figures 1 K and 1L), suggesting that increased running of KO mice is not due to trophic actions associated with leptin. Finally, in view of the link between LepR signaling in DA neurons and anxiety (), we verified anxiety-like behavior. Exploration in the elevated-plus maze and open field was similar between genotypes ( Figures S4 A–S4D). Thus, loss of STAT3 in DA neurons enhances spontaneous activity and endurance running in a manner unrelated to anxiety, results that coincide with observations that VTA leptin inhibits wheel running in a rat model of anorexia-induced hyperactivity (). Hyperactivity and motor restlessness is a detrimental clinical feature of anorexia nervosa that has been tied to hypoleptinemia (), and thus these findings have implications for understanding anorexia-induced hyperactivity.

STAT3 mice in which loxP sites flank exon 22 of the STAT3 gene that encodes tyrosine 705 that is essential for STAT3 activation () were bred with mice that express Cre recombinase under the control of the dopamine transporter (DAT). STAT3;DATmice (“STAT3”) had similar weaning weights ( Figure 1 A, inset). The recombined STAT3 gene was observed in tissue in which DAT is expressed ( Figure S1 A) and was specific to DA neurons ( Figure S1 B).

Discussion

The present study reveals leptin as an important signal controlling the rewarding impact of running and identifies a key role for STAT3 signaling in DA neurons in this process. STAT3 deletion led to substantial increases in spontaneous activity and endurance running, whereas targeted viral-mediated restoration of STAT3 reversed this effect. STAT3 loss-of-function had little influence on the anorectic actions of leptin, hedonic, or compulsive feeding behavior, results that highlight the specific involvement of STAT3 signaling in DA neurons in the control of physical activity. By tying behavioral changes to decreases in DA overflow and reduced mesolimbic function, the results underscore STAT3 as an important regulator of mesolimbic DA tone, physical activity, and the motivational effects of running.

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Flier J.S. Leptin regulation of the mesoaccumbens dopamine pathway. Few studies have investigated the mechanisms underlying the effects of leptin on physical activity. The hypoactivity of obese, LepR-deficient mice is corrected by restoration of LepR to the mediobasal hypothalamus () or proopiomelanocortin neurons (), underscoring the role of the hypothalamus in the actions of leptin to stimulate locomotor behavior. However, leptin suppression of physical activity when food is limited suggests the presence of distinct neural mechanisms. We speculate that in conditions of restricted food availability the mesolimbic DA system engages motivational processes concerned with obtaining food and more readily responds to leptin to decrease appetitive physical activity. On the other hand, during fed states, the actions of leptin may be biased toward hypothalamic processes that could increase physical activity as a means to maintain energy homeostasis. Our results point to reduced LepR-STAT3 signaling in midbrain DA neurons as an important mechanism mediating increased physical activity by food restriction and falling leptin levels. Consideration must be given to the fact that signals other than leptin activate STAT3, and thus we cannot rule out that the behavioral and biochemical effects observed could be independent of leptin. Our findings that VTA leptin decreases the rewarding effects of running in a STAT3-dependent manner suggest that leptin is involved; however, the concentrations of leptin administered may not represent physiological levels found in the VTA. In spite of this, a previous study noted that phosphorylation of STAT3 at Tyr705 was not evident in the VTA of LepR-deficient mice () to suggest that other signals (e.g., interleukin-6) are not activating STAT3 in DA neurons in basal conditions.

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Stoving R.K. Exercise Addiction in Men Is Associated With Lower Fat-Adjusted Leptin Levels. In conditions where food is abundant and easy to access, the runner’s high could prove beneficial for encouraging physical activity. It is compelling that chronic, but not acute, exercise lowers leptin levels in men (), and so endurance training could be intrinsically rewarding by moderating LepR-STAT3 signaling in DA neurons. The rewarding properties of physical activity can foster excessive exercise and compulsive behavior characteristic of addiction in some individuals. Intriguingly, a recent report shows that exercise addiction in men is associated with low, fat-adjusted leptin levels (). It remains to be elucidated if genetic or physiological factors could contribute to a larger drop in leptin or enhanced LepR sensitivity in response to food restriction or exercise during normal or pathological (e.g., anorexia) states that would increase the propensity for physical activity.