Intracerebroventricular Leptin Infusion Research Articles

Chronic central nervous system leptin administration attenuates kidney dysfunction and injury in a model of ischemia/reperfusion-induced acute kidney injury.

In the present study we examined whether chronic intracerebroventricular (ICV) leptin administration protects against ischemia/reperfusion (I/R)-induced acute kidney injury (AKI). Twelve-week-old male rats were implanted with an ICV cannula into the right lateral ventricle and 8-10 days after surgery, leptin (0.021 mg/hr, n=8) or saline vehicle (0.5 ml/h, n=8) was infused via osmotic minipump connected to the ICV cannula for 12 days. On day 8 of leptin or vehicle infusion, rats were submitted to unilateral ischemia/reperfusion (UIR) by clamping the left pedicle for 30 min. To control for leptin-induced reductions in food intake, the vehicle-treated group was pair-fed (UIR-PF) to match the same amount of food consumed by leptin-treated (UIR-Leptin) rats. On the 12th day of leptin or vehicle infusion (4th day after AKI), single-left kidney glomerular filtration rate (GFR) was measured, blood samples were collected to quantify white blood cells, and kidneys were collected for histological assessment of injury. UIR-Leptin treated rats showed reduced right and left kidney weights (right: 1040±24 vs. 1281±36 mg; left: 1127±71 vs. 1707±45 mg, for UIR-Leptin and UIR-PF, respectively). ICV leptin infusion improved GFR (0.50±0.06 vs. 0.13±0.03 ml/min/g KW) and reduced kidney injury scores. ICV leptin treatment also attenuated the reduction in circulating adiponectin levels that was observed in UIR-PF rats, and increased circulating white blood cells count compared to UIR-PF rats (16.3±1.3 vs. 9.8±0.6 k/mL). Therefore, we show that leptin, via its actions on the central nervous system, confers significant protection against major kidney dysfunction and injury in a model of ischemia/reperfusion-induced AKI.

Setmelanotide Administration Improves Cardiac Systolic Function in Sirtuin-3 Deficient Mice

We previously showed that activation of the brain leptin-melanocortin system, particularly melanocortin-4 receptors (MC4R), markedly improves cardiac function in experimental models of heart failure (HF) via mechanisms that are still poorly understood. Sirtuin 3 (Sirt3) is an important regulator of mitochondrial oxidative phosphorylation and deficiency of Sirt3 has been implicated in cardiovascular diseases. We recently found increased Sirt3 expression as well as increased levels of NAD+, the main Sirt3 activator, in cardiomyocytes of rats treated with chronic intracerebroventricular infusion of leptin at a dose that does not spill over into the systemic circulation. Therefore, we hypothesized that stimulation of Sirt3 activity is required for the improvement of cardiac function during activation of the brain leptin-MC4R axis. To test this hypothesis, we first examined if chronic MC4R activation, using an MC4R agonist, setmelanotide, improves cardiac function in naïve Sirt3-deficient (Sirt3-KO) mice. Baseline cardiac function was assessed by echocardiography (VisualSonics-VEVO3100®) in male and female Sirt3-KO and wild-type (WT) control mice (n=5/group) at 20 weeks of age, and then weekly during setmelanotide treatment (2 mg/kg/day — i.p. via osmotic minipump model 1002) for 3 weeks. Chronic MC4R activation with setmelanotide transiently reduced food intake in both groups and promoted weight loss in Sirt3-KO mice. Setmelanotide treatment improved systolic function in WT mice as evidenced by increases in ejection fraction and fractional shortening (EF: 52.8±4.4 vs. 72.4±4.4%; FS: 26.9±2.7 vs. 42.8±6.4%, for baseline vs. week 3, respectively) and other measures of systolic function including circumferential and longitudinal strains. Despite showing a tendency for reduced systolic function compared to WT controls at baseline, Sirt3-KO mice also exhibited improvement in systolic function with setmelanotide administration (EF: 44.1±1.8 vs. 65.3±2.9% and FS: 21.4±1.0 vs. 35.1±2.1%). No major changes were observed in measures of diastolic function. These results suggest that in a non-HF model, MC4R activation does not require stimulation of Sirt3 to improve indices of systolic function. (1R01HL163076, 1R01DK121411, P20GM104357, P30GM149404, U54GM115428, and AHA-835218). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Abstract 14283: Brain-Mediated Cardioprotective Actions of Leptin: Potential Role of Brown Adipose Tissue and Its Release of Extracellular Vesicles

Background: We recently demonstrated that leptin has powerful central nervous system (CNS) actions that dramatically improve cardiac function following ischemia/reperfusion (IR) injury. How the brain communicates with the heart during CNS leptin infusion to improve cardiac function is unknown. One potential mechanism is activation of brown adipose tissue (BAT) which may be an important source of extracellular vesicles (EVs) carrying cardioprotective microRNAs (miR), including miR-125b-5p. Hypothesis: We investigated whether BAT contributes to the cardioprotective effect of CNS leptin infusion and the potential role of EVs carrying the miR-125b-5p in this protection. Methods: Male Sprague-Dawley rats were submitted to interscapular BAT ablation (iBATx) and instrumented with an intracerebroventricular (ICV) cannula in the brain lateral ventricle. After recovery and baseline assessment of cardiac function by echocardiography (ECHO), myocardial IR was induced by temporary ligation of the left anterior descending coronary artery. Vehicle (saline, 0.5 μL/hr) or leptin (0.62 μg/hr) was infused ICV chronically for 28 days using osmotic pumps. Cardiac function was assessed weekly by ECHO. Another set of animals with intact iBAT was treated with ICV leptin or vehicle for 4 weeks and had their iBAT collected for EVs isolation and quantification. Expression of the miR-125b-5p was measured in the heart of leptin treated rats with intact iBAT or iBATx. Results: Compared to vehicle-treated animals, ICV leptin infusion increased iBAT-derived EVs and removal of iBAT prevented leptin’s CNS-mediated cardioprotective effects after IR injury as indicated by a lack of improvement in ejection fraction (26±6 vs 41±3%) and stroke volume (162±31 vs 275±19μL) at week 4 post-IR. Expression of the mature miR-125b-5p was increased in the heart of ICV leptin treated rats and iBATx abolished this effect. Conclusion: These results indicate that the BAT plays an important role in mediating the cardioprotective effect of CNS leptin administration after IR injury and suggest that iBAT-derived EVs may be involved in delivering the miR-125b-5p to the heart during central leptin infusion. AHA 835218, 1R01HL163076, P20GM104357, and P20GM104320-07

Central Nervous System Actions of Leptin Improve Cardiac Function After Ischemia-Reperfusion: Roles of Sympathetic Innervation and Sex Differences.

Background Therapeutic strategies for preventing paradoxical reperfusion injury after myocardial ischemia are limited. We tested whether central nervous system actions of leptin induce important protective effects on cardiac function and metabolism after myocardial ischemia/reperfusion (I/R) injury, the role of cardiac sympathetic innervation in mediating these effects, and whether there are major sex differences in the cardioprotective effects of chronic central nervous system leptin infusion. Methods and Results Myocardial I/R was induced by temporary ligation of the left descending coronary artery in male and female Wistar rats instrumented with intracerebroventricular cannula in the lateral ventricle. Vehicle or leptin (0.62 μg/h) infusion was started immediately after reperfusion and continued for 28 days using osmotic minipumps connected to the intracerebroventricular cannula. Cardiac function was assessed by echocardiography, ventricular pressures, and exercise performance. Intracerebroventricular leptin treatment markedly attenuated cardiac dysfunction post-I/R as evidenced by improved ejection fraction (56.7±1.9 versus 22.6%±1.1%), maximal rate of left ventricle rise (11 680±2122 versus 5022±441 mm Hg) and exercise performance (-4.2±7.9 versus -68.2±3.8 Δ%) compared with vehicle-treated rats. Intracerebroventricular leptin infusion reduced infarct size in females, but not males, when compared with ad-lib fed or pair-fed saline-treated rats. Intracerebroventricular leptin treatment also increased cardiac NAD+/NADH content (≈10-fold) and improved mitochondrial function when compared with vehicle treatment. Cervical ganglia denervation did not attenuate the cardiac protective effects of leptin after I/R injury. Conclusions These data indicate that leptin, via its central nervous system actions, markedly improves overall heart function and mitochondrial metabolism after I/R injury regardless of sex, effects that are largely independent of cardiac sympathetic innervation.

Chronic Central Nervous System Leptin Infusion Improves Cardiac Function and Metabolism after Ischemia/Reperfusion Injury

Important adverse changes in cardiac energy metabolism occur after ischemia/reperfusion injury (I/R), contributing to the worsening of cardiac function and development of heart failure. We recently showed that leptin, via its actions on the central nervous system (CNS), improves left ventricular (LV) function in a model of heart failure induced by permanent ligation of the left anterior descending coronary artery (LAD). In the present study, we examined whether the CNS‐ effects of leptin protect against myocardial I/R injury, and if chronic intracerebroventricular (ICV) leptin infusion improves cardiac substrate utilization, assessed by markers of myocardial fatty acid (FA) and glucose oxidation, NAD+/NADH redox state and plasma levels of β‐hydroxybutyrate (β‐HOB). Male Wistar rats were instrumented with an ICV cannula in the brain lateral ventricle. After recovery and baseline assessment of cardiac function by echocardiography (ECHO), myocardial I/R was induced by temporary (60 min) ligation of the LAD. Vehicle (saline, 0.5 µL/hr) or leptin (0.62 µg/hr) was infused chronically for 28 days starting 20 min after reperfusion using osmotic minipumps connected to the ICV cannula. ECHO assessment of cardiac function was performed weekly. At the end of week 4, +dP/dtmax was accessed by LV catheterization. Hearts and plasma samples were collected for evaluation of CD36, PPAR‐δ, PDK4 and p‐PDH1/PDH1 by western blot and cardiac NAD+/NADH ratio and plasma levels of β‐OHB were measured by ELISA. Our results showed that ICV leptin treatment improved cardiac function as evidenced by increased ejection fraction 4 weeks after I/R (46±3 vs. 26±3 %) and +dP/dtmax (10387±1686 vs. 5022±442 mmHg/s) when compared with vehicle‐treated rats. ICV leptin infusion also significantly increased cardiac protein expression of CD36 (1.4±0.1 vs. 1.0±0.1 au), PPAR‐δ (1.8±0.1vs. 1.0±0.01 au), PDK4 (1.8±0.1 vs. 1.0±0.1 au) and p‐PDH1/PDH1 (2.2±0.3 vs. 1.0±0.2 au) when compared with vehicle‐treated animals. In addition, ICV leptin infusion increased cardiac NAD+/NADH ratio (6.8±2 vs. 1.02±0.3 pmol/µL) and reduced plasma levels of β‐OHB (18.2±3 vs. 62.8±7.0 nmol/µL). These results demonstrate that chronic ICV leptin infusion improves cardiac function following I/R injury and suggest that leptin’s CNS‐mediated cardioprotective effects may involve improved myocardial FA oxidation and NAD+/NADH redox state, reducing the reliance on ketone bodies (β‐OHB) as an energy source.

Abstract P321: Chronic Central Nervous System Leptin Infusion Increases Cardiomyocyte Mitochondrial Function And Biogenesis And Improves Cardiac Function After Ischemia-reperfusion Injury.

Mitochondrial (MT) dysfunction plays an important role in the pathophysiology of myocardial ischemia/reperfusion injury (I/R). We recently showed that leptin, via its actions in the central nervous system (CNS), improves left ventricular (LV) function in a model of heart failure induced by permanent ligation of the left anterior descending coronary artery (LAD). In the present study, we examined if the CNS effects of leptin protect against myocardial ischemia/reperfusion (I/R) injury, and whether chronic intracerebroventricular (ICV) leptin infusion increases MT function and biogenesis in the non-infarcted area of the LV that is at risk but still viable. Male Wistar rats were instrumented with an ICV cannula in the brain lateral ventricle. After recovery and baseline assessment of cardiac function by echocardiography (ECHO), myocardial I/R was induced by temporary (60 min) ligation of the LAD. Vehicle (saline, 0.5 μL/hr) or leptin (0.62 μg/day) was infused chronically for 28 days starting 20 min after reperfusion using osmotic minipumps connected to the ICV cannula. ECHO assessment of cardiac function was performed every week. At the end of week 4, +dP/dt max and Tau were accessed by LV catheterization. Hearts were then collected for evaluation of MT function in isolated cardiac fibers using Oroboros oxygraphy-2k respirometer. ICV leptin treatment improved systolic and diastolic function as evidenced by increased ejection fraction 4 weeks after I/R (46±3 vs. 26±3 %), stroke volume (353±19 vs. 193±27 μL), +dP/dt max (10387±1686 vs. 5022±442 mmHg/s) and reduced Tau (6.5±0.3 vs. 8±0.3 ms) when compared with vehicle-treated rats. In addition, ICV leptin infusion significantly increased ATP-linked respiration (55±3 vs. 39±1 %), reduced proton leak (45±3 vs. 61±1 %), and improved MT reserve capacity (42±5 vs. 27±2 %). Improved MT function was associated with increased MT Complex I- mediated respiration (47±6 vs. 26±1 pmolO 2 /s - mg). ICV leptin treatment also increased PGC1 α protein expression and COX3 gene expression, indicating enhanced MT biogenesis. These results demonstrate that chronic ICV leptin infusion improves cardiac function following I/R injury and suggests that leptin’s CNS-mediated cardioprotective effects may involve improved myocardial MT function and biogenesis.

Chronic Antidiabetic Actions of Leptin: Evidence From Parabiosis Studies for a CNS-Derived Circulating Antidiabetic Factor.

We used parabiosis to determine whether the central nervous system (CNS)-mediated antidiabetic effects of leptin are mediated by release of brain-derived circulating factors. Parabiosis was surgically induced at 4 weeks of age, and an intracerebroventricular (ICV) cannula was placed in the lateral cerebral ventricle at 12 weeks of age for ICV infusion of leptin or saline vehicle. Ten days after surgery, food intake, body weight, and blood glucose were measured for 5 consecutive days, and insulin-deficiency diabetes was induced in all rats by a single streptozotocin (STZ) injection (40 mg/kg). Five days after STZ injection, leptin or vehicle was infused ICV for 7 days, followed by 5-day recovery period. STZ increased blood glucose and food intake. Chronic ICV leptin infusion restored normoglycemia in leptin-infused rats while reducing blood glucose by ∼27% in conjoined vehicle-infused rats. This glucose reduction was caused mainly by decreased hepatic gluconeogenesis. Chronic ICV leptin infusion also reduced net cumulative food intake and increased GLUT4 expression in skeletal muscle in leptin/vehicle compared with vehicle/vehicle conjoined rats. These results indicate that leptin's CNS-mediated antidiabetic effects are mediated, in part, by release into the systemic circulation of leptin-stimulated factors that enhance glucose utilization and reduce liver gluconeogenesis.

Effects of intracerebroventricular leptin and orexin-A on the baroreflex control of renal sympathetic nerve activity in conscious rats fed a normal or high-fat diet.

This study examined the effect of leptin and orexin-A on autonomic baroreflex control in conscious Wistar rats exposed to high-fat (45% fat) or normal (3.4%) diet for 4weeks. Renal sympathetic nerve activity (RSNA), mean arterial pressure (MAP) and heart rate (HR) were monitored during the generation of baroreflex gain curves and acute volume expansion (VEP). Intracerebroventricular (ICV) leptin (1μg/min) increased RSNA in the normal diet group (0.31±0.04 vs 0.23±0.03mV/s) and MAP in the high-fat diet group (115±5 vs 105±5mm Hg, P<.05). Orexin-A (50ng/min) increased RSNA, HR and MAP in the high-fat diet group (0.26±0.03 vs 0.22±0.02mV/s, 454±8 vs 417±12beats/min, 117±1 vs 108±1mm Hg) and the normal diet group (0.18±0.05 vs 0.17±0.05mV/s, 465±10 vs 426±6beats/min, 116±2 vs 104±3 mmHg). Baroreflex sensitivity for RSNA was increased during ICV leptin by 50% in the normal diet group, compared to 14% in the high-fat diet group (P<.05). Similarly, orexin-A increased baroreflex sensitivity by 56% and 50% in the high-fat and normal diet groups, respectively (all P<.05). During ICV saline, VEP decreased RSNA by 31±5% (P<.05) after 10minutes and the magnitude of this response was blunted during ICV infusion of leptin (17±2%, P<.05) but not orexin-A in the normal diet group. RSNA response to VEP was not changed during ICV leptin or orexin-A in the high-fat diet group. These findings indicate possible central roles for leptin and orexin-A in modulating the baroreflexes under normal or increased fat intake in conscious rats and potential therapeutic approaches for obesity associated hypertension.

Abstract 16200: Chronic Central Nervous System Actions of Leptin Protect Against Myocardial Ischemia/reperfusion Injury

While myocardial reperfusion is the most effective therapy to reduce mortality after myocardial infarction, it can paradoxically exacerbate ischemic injury. We recently showed that leptin, via its actions on the central nervous system (CNS), improved left ventricular function in a model of heart failure induced by permanent ligation of the left anterior coronary artery (LAD). To investigate whether leptin also protects the heart against ischemia/reperfusion (I/R) injury via its actions in the CNS, we instrumented female Wistar rats (15 weeks of age) with an intracerebroventricular (ICV) cannula into the lateral ventricle, and after 7 days of recovery and baseline assessment of cardiac function by echocardiography (VisualSonics VEVO-3100®), myocardial I/R was induced by temporary LAD ligation (60 min). Vehicle (saline, 0.5 μL/hr, n=6) or leptin (15 μg/day, n=7) were infused ICV for 28 consecutive days starting 20 min after reperfusion using osmotic minipump connected to the ICV cannula. Echocardiographic assessment of cardiac function was performed every week and at the end of the 4 th week of treatment, the heart was collected and processed for protein analysis and histological evaluation of infarct size and collagen deposition. Compared to vehicle treatment, chronic ICV leptin infusion significantly reduced infarcted area (21±2 vs. 37±4 %), septal collagen deposition (2.2±0.2 vs. 4.0±0.7 %), and markedly attenuated systolic dysfunction as evidenced by increased ejection fraction 4 weeks post I/R (59±1 vs. 30±2%), stroke volume (296±19 vs. 159±8 μL) and cardiac output (108±5 vs.63±4 μL/min). ICV leptin infusion also prevented the increase in left atrium to aorta diameter ratio (1.5±0.1 vs. 2±0.1 mm), an index of cardiac congestion. In addition, we found a 2-fold increase in the ratio of phospho-acetyl-CoA carboxylase (p-ACC) to total ACC protein expression, a marker of fatty acid oxidation, in hearts of leptin-treated rats compared with vehicle infusion. These results indicate that leptin exerts powerful beneficial CNS-mediated effects on the heart that improve systolic function and protect the myocardium against I/R injury. Our results also suggest that these beneficial actions may involve improved myocardial bioenergetics.

Chronic Intracerebroventricular Leptin Infusion Attenuates Cardiac Dysfunction After Myocardial Infarction

Leptin is an important regulator of energy balance and substrate utilization in various tissues including the heart. In the present study we tested whether activation of leptin receptors in the central nervous system (CNS) confers protection against cardiac dysfunction after myocardial infarction (MI) induced by permanent ligation of the left anterior descending coronary artery (LAD). Male Sprague‐Dawley rats at 12 weeks of age were implanted with BP telemetry transmitters and an intracerebroventricular (ICV) cannula was inserted into the lateral ventricle. After 10 days of recovery, mean arterial pressure (MAP) and heart rate (HR) were measured 24‐hrs/day by telemetry and cardiac function was assessed by echocardiography (VisualSonics‐VEVO3100®). After stable baseline measurements for 4 days, the LAD was permanently ligated and saline vehicle (0.5 μL/hr, n=6) or leptin (15 μg/day, n=7) were infused ICV via osmotic minipump for 28 consecutive days. Compared to vehicle treatment, chronic ICV leptin infusion for 4 weeks markedly attenuated cardiac dysfunction caused by MI as evidenced by normalization of global cardiac radial strain (39±4 to 36±5 vs. 37±4 to 25±3 %), cardiac output (92±5 to 110±10 vs. 105±6 to 83±9 ml/min) and left atrium to aorta ratio, an index of cardiac congestion, (1.6±0.1 to 1.6±0.1 vs. 1.7±0.1 to 2.3±0.1). Central leptin administration also significantly improved ejection fraction (from 35±3 to 49±2 %) and fractional shortening (from 12.4±0.8 to 18.6±1.4 %, for week 1 and week 4 post‐MI, respectively), whereas no improvement was observed in vehicle‐treated rats (from 36±4 to 30±2 %, and from 12.2±1.7 to 10.1±1.3 %, respectively). In addition, ICV leptin infusion prevented the bradycardia observed in vehicle‐treated rats (354±3 to 370±6 vs. 360±7 to 323±6 bpm) but did not attenuate the reduction in MAP evoked by MI (101±4 to 94±3 vs. 102±3 to 97±2 mmHg). There was no difference in infarcted area between groups (30±3 vs. 29±5 %). These results indicate that leptin can improve cardiac function and attenuate progression of heart failure after MI via a CNS‐mediated mechanism.Support or Funding InformationNHLBI PO1HL51971, NIGMS P20GM104357 and NIGMS U54GM115428This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Differential Regulation of Cardiac Substrate Utilization in Response to Chronic Central Nervous System Administration of Leptin and Melanotan II in Rats with Myocardial Infarction

The brain leptin‐melanocortin system pathway plays an important role in regulating many physiological functions including energy homeostasis. We found that chronic intracerebroventricular (ICV) infusion of leptin or the melanocortin 4 receptor (MC4R) agonist Melanotan II, MTII, markedly attenuated cardiac dysfunction in rats with myocardial infarction (MI) induced by permanent left anterior descending coronary artery (LAD) ligation. In the present study we examined if activation of leptin‐MC4R axis in the central nervous system (CNS) alters cardiac substrate utilization in rats with MI. Male Sprague‐Dawley rats at 15 weeks of age were implanted with an ICV cannula into the lateral ventricle. After 10 days of recovery, the LAD was permanently ligated and saline vehicle (0.5 μL/hr, n=5), leptin (15 μg/day, n=5) or MTII (240 ng/day, n=3) was infused ICV via osmotic minipump for 14 consecutive days. On day 14 of treatment the hearts were quickly excised and perfused ex vivo in the working mode with glucose (5.5 mmol/L) and the fatty acid oleate (0.4 mmol/L). After a 5‐min baseline equilibration period the hearts were perfused for an additional 20 min and coronary flow samples were collected every 5 min to determine rates of substrate oxidation by quantitative measurement of [14C]O2 and [3H]2O using [U‐14C]glucose (0.08 μCi/ml) and [9,10‐3H]oleate (0.1 μCi/ml) as the radiolabeled tracers. Compared to vehicle treatment, chronic ICV leptin infusion for 2 weeks markedly increased glucose oxidation (0.60±0.04 vs. 0.27±0.02 μmol/min/g dry heart weight) without altering oleate oxidation (0.95±0.01 vs. 0.90±0.02 μmol/min/g dry heart weight). Chronic central MTII infusion did not alter glucose oxidation (0.25±0.03 vs. 0.27±0.02 μmol/min/g dry heart weight) but increased fatty acid oxidation compared to vehicle treatment (1.13±0.01 vs. 0.90±0.02 μmol/min/g dry heart weight). These results suggest the CNSmechanisms by which leptin and MC4R activation improve cardiac function post MI may involve increased myocardial bioenergetics by differential modulation of glucose and fatty acid utilization.Support or Funding InformationNHLBI PO1HL51971, NIGMS P20GM104357 and NIGMS U54GM115428This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Evidence for a circulating factor released by the brain that contributes to chronic antidiabetic actions of leptin

We previously demonstrated that leptin has powerful central nervous system (CNS)‐mediated antidiabetic actions that are independent of the autonomic nervous system (ANS) and the pituitary. In this study, we examined if a circulating factor released by the brain contributes to leptin's chronic antidiabetic actions in insulin deficient rats. Male inbred 4‐week‐old Lewis rats (n=14) were conjoined by attaching their skin to allow shared circulation. Six weeks later, the rats were implanted with an intracerebroventricular (ICV) cannula into the lateral ventricle to infuse leptin in one rat and vehicle in the other rat of each pair. Eight days after surgery, food intake, body weight and blood glucose concentration were measured for 5 consecutive days and then streptozotocin (STZ) was injected (40 mg/kg) to induce insulin‐deficient diabetes in both rats. Five days after STZ injection, leptin (15 mg/day) or vehicle (0.5 ml/h) was delivered ICV continuously for 7 days via osmotic minipumps. STZ injection significantly increased average blood glucose concentration (359±19 vs. 79±6 mg/dL) and food intake (49±2 vs. 39±3 g/day). Despite higher food intake, average body weight decreased after STZ injection (619±23 vs. 664±23 g). Chronic ICV leptin infusion restored normoglycemia (110±31 vs. 359±19 mg/dL) in leptin‐treated rats and reduced blood glucose by ~27% in the paired parabiotic rat infused with vehicle (269±20 vs. 366±41 mg/dL). Leptin treatment did not reduce food intake (53±20 vs. 49±23 g/day) or body weight (619±23 vs. 610±20 g) in the paired rats. After stopping leptin treatment, blood glucose concentration returned to diabetic values in leptin‐treated (305±44 vs. 110±31 mg/dL) as well as in vehicle‐treated rats (330±11 vs. 269±20 mg/dL), and food intake increased 37% (67±5 vs. 49±3 g/day). These results suggest that the chronic antidiabetic effects of leptin may involve release into the systemic circulation of a brain‐derived neuroendocrine factor that enhances glucose homeostasis in insulin deficient rats. (NHLBI‐PO1HL51971, NIGMS P20GM104357)Support or Funding InformationDepartment of Physiology and Biophysics and Mississippi Center for Obesity Research. University of Mississippi Medical Center.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Abstract P391: Local Ionotropic Glutamate Receptors Contribute to the Sympathoexcitatory Effects of Leptin in the Paraventricular Nucleus

In male rats, intracerebroventricular infusion of leptin increases sympathetic nerve activity (SNA), and this response can be partially reversed by nanoinjections of the ionotropic glutamate receptor (iGluR) blocker, kynurenate (KYN), and also the melanocortin type 3/4 receptor (MC3/4R) antagonist, SHU9119, into the hypothalamic paraventricular nucleus (PVN). These results suggest that leptin increases SNA in part via activation of PVN iGluR and MC3/4R. More recently, we found that injection of leptin directly into the PVN also dose-dependently increases SNA. However, the role of PVN iGluR or MC3/4R in this response is unknown. Therefore, we determined if PVN KYN (2.7 nmol in 60 nL) or SHU9119 (30 pmol) reverses the increases in lumbar SNA (LSNA), mean arterial pressure (MAP), and heart rate (HR) evoked by bilateral PVN nanoinjections of leptin (30 ng in 60 nL; n=6) or, as a control, of artificial cerebrospinal fluid (aCSF; n=4). After ninety min, PVN leptin increased (all P&lt;0.05) LSNA to 164±10 % of control, MAP from 106±5 to 118±5 mmHg, and HR from 360±12 to 399±16 bpm. Then, PVN injections of KYN decreased (all P&lt;0.05) LSNA to 133±8 % control, MAP to 108±4 mmHg, and HR to 379±16 bpm. On the other hand, neither PVN aCSF injections (LSNA, to 105±3 % control; MAP, from 107±8 to 109±7 mmHg; and HR, from 363±31 to 367±20 bpm) nor subsequent PVN KYN injections (LSNA, to 102±5 % control; MAP, to 108±6 mmHg; and HR, to 373±17 bpm) significantly altered these variables. In a second set of animals, while PVN leptin (n=4) elicited similar increases (all P&lt;0.05) in LSNA (to 158±8 % control), MAP (from 101±6 to 113±4 mmHg), and HR (from 367±19 to 405±11 bpm), subsequent SHU9119 had no effects (LSNA, to 151±7 % control; MAP, to 108±5 mmHg; and HR, to 400±14 bpm). Again, no changes in these variables were observed in response to PVN injections of aCSF (n=4), even when followed by SHU9119 (LSNA, to 114±11 and then 110±13 % control; MAP, from 107±7 to 109±8 and then 106±9 mmHg; and HR, 357±31 to 355±24 and then 354± 25 bpm). In conclusion, activation of local iGluR, but not MC3/4R, contribute to the increases in LSNA, MAP, and HR in response to PVN leptin.

Chronic Central Leptin Infusion Improves Cardiac Function in Diabetic Rats With Heart Failure

Myocardial infarction (MI) is a major cause of mortality and morbity and the leading cause of heart failure (HF) worldwide. We previously demonstrated that leptin, a peptide hormone produced by adipocytes, exerts powerful antidiabetic effects via activation of its receptors in the central nervous system (CNS). We also showed that chronic central leptin infusion restores intrinsic heart rate, cardiac sympathetic‐vagal balance and baroreflex sensitivity in uncontrolled diabetes. Therefore, in this study we examined if chronic intracerebroventricular infusion of leptin improves cardiac function in rats with HF induced by MI. Wistar Munich rats (275–300 g, n=4–5/group) were submitted to MI by ligation of the left descending coronary artery. Diabetes was induced one week later by a single injection of streptozotocin (STZ, 35 mg/kg, i.p.). Seven days later, rats were instrumented with an intracerebroventricular (ICV) cannula into the lateral ventricle for continuous infusion of leptin (0.62 μg/h, 0.5 μL/h, n=5) or vehicle (0.5 μL/h, n=4), for 4 weeks using osmotic pumps (Alzet, model 2002). Cardiac function was evaluated 6 days after MI surgery and on days 14 and 28 of ICV leptin or vehicle infusion using the echocardiograph VEVO 2100 (VisualSonics, Canada). STZ caused severe hyperglycemia in both groups (ranging from 472±17 to 477±14 mg/dl) which was completely normalized by chronic ICV leptin infusion (120±8 mg/dl). Compared to vehicle group, leptin infusion for 4 weeks also markedly ameliorated cardiac function as evidenced by lower left atrium/aorta ratio (1.73±0.07 vs. 2.63±0.23 mm) and higher cardiac output (113.8±27.6 vs. 69.5±10.0 ml/min) and ejection fraction (36.7±2.3 vs. 30.7±8.7 %). These results suggest that in addition to its powerful CNS‐mediated antidiabetic effects, leptin attenuates the progression of HF after MI in diabetic rats by improving cardiac function.Support or Funding InformationNHLBI‐PO1HL51971, NIGMS‐P20GM104357, FAPESP, CNPq

Role of autonomic nervous system in chronic CNS-mediated antidiabetic action of leptin.

This study tested whether ganglionic blockade or hepatic vagotomy attenuates the chronic central nervous system (CNS)-mediated antidiabetic and cardiovascular effects of leptin. Male Sprague-Dawley rats were instrumented with telemetry probes and arterial and venous catheters for determination of blood pressure (BP), heart rate (HR), blood sampling, and intravenous (iv) infusions. An intracerebroventricular (ICV) cannula was placed into the brain lateral ventricle for infusion of leptin or vehicle. After control measurements, streptozotocin (STZ) was injected iv (50 mg/kg) to induce diabetes, and 5 days later leptin (n = 6) or saline vehicle (n = 5) was infused ICV for 12 days via osmotic pumps. Beginning on day 6 of leptin treatment, the ganglionic blocker hexamethonium (15 mg·kg-1·day-1 iv) was infused, while leptin infusion was continued, to assess the role of the autonomic nervous system. Induction of diabetes was associated with increases in blood glucose (98 ± 7 to 350 ± 19 mg/dl), food intake (23 ± 3 to 43 ± 3 g/day), decreases in HR (-70 ± 11 beats/min), polyuria, and increased water consumption, which were all completely normalized by ICV leptin infusion. Although hexamethonium attenuated leptin's effect on HR, it failed to impair leptin's ability to restore euglycemia or to prevent the polyuria or increased water intake in STZ-diabetic rats. We also found that after pretreatment with hexamethonium (n = 8), ICV leptin infusion, during continued ganglionic blockade, completely normalized blood glucose in diabetic rats. In addition, selective hepatic vagotomy did not attenuate leptin's ability to restore euglycemia in diabetic rats. These results suggest that leptin's powerful chronic CNS antidiabetic actions are mediated primarily via nonautonomic mechanisms.

Evidence against hypothalamic-pituitary-adrenal axis suppression in the antidiabetic action of leptin.

Leptin administration restores euglycemia in rodents with severe insulin-deficient diabetes, and recent studies to explain this phenomenon have focused on the ability of leptin to normalize excessive hypothalamic-pituitary-adrenal (HPA) axis activity. Here, we employed a streptozotocin-induced rat model (STZ-DM) of uncontrolled insulin-deficient diabetes mellitus (uDM) to investigate the contribution of HPA axis suppression to leptin-mediated glucose lowering. Specifically, we asked if HPA axis activation is required for diabetic hyperglycemia, whether HPA axis normalization can be achieved using a dose of leptin below that needed to normalize glycemia, and if the ability of leptin to lower plasma glucocorticoid levels is required for its antidiabetic action. In STZ-DM rats, neither adrenalectomy-induced (ADX-induced) glucocorticoid deficiency nor pharmacological glucocorticoid receptor blockade lowered elevated blood glucose levels. Although elevated plasma levels of corticosterone were normalized by i.v. leptin infusion at a dose that raises low plasma levels into the physiological range, diabetic hyperglycemia was not altered. Lastly, the potent glucose-lowering effect of continuous intracerebroventricular leptin infusion was not impacted by systemic administration of corticosterone at a dose that maintained elevated plasma levels characteristic of STZ-DM. We conclude that, although restoring low plasma leptin levels into the physiological range effectively normalizes increased HPA axis activity in rats with uDM, this effect is neither necessary nor sufficient to explain leptin's antidiabetic action.

Differences in mitochondrial DNA inheritance and function align with body conformation in genetically lean and fat sheep.

Body weight and adiposity are determined by the balance between energy intake, energy expenditure, and nutrient deposition. We have identified differences in appetite-regulating peptides in sheep selectively bred to be either lean or fat, wherein gene expression for orexin and melanin-concentrating hormone are elevated in the lean group. Despite this, the underlying mechanisms leading to differences in body composition in the lean and fat lines remains unknown. We measured postprandial temperature in adipose tissue and muscle to ascertain whether a difference in thermogenesis is associated with the difference in body composition in genetically lean (n = 8) and fat (n = 12) ewes. Body weight was higher (P < 0.01) but percent fat mass was lower (P < 0.001) in the lean group. The percent lean mass was similar in lean and fat groups. Animals received intracerebroventricular cannulae and temperature probes implanted into the retroperitoneal fat and the hind-limb skeletal muscle (vastus lateralis). Animals were meal fed (1100-1600 h) to entrain postprandial thermogenesis. Food intake was similar between lean and fat animals. Postprandial thermogenesis was greater (P < 0.05) in the retroperitoneal adipose tissue of lean animals but not in skeletal muscle. Intracerebroventricular infusion of leptin reduced (P< 0.05) food intake by an equal extent in both groups. Postprandial expression of UCP1 mRNA was greater (P < 0.05) in retroperitoneal fat of lean animals, with similar UCP3 expression in skeletal muscle. Mitochondrial genome sequencing indicated haplotypic clustering in lean and fat animals within both the encoding and nonencoding regions. This demonstrates that differences in body composition may be underpinned by differences in thermogenesis, specifically within adipose tissue. Furthermore, thermogenic differences may be associated with specific mitochondrial DNA haplotypes, suggesting a strong genetic component inherited through the maternal lineage.

Age versus nutritional state in the development of central leptin resistance

Leptin, a catabolic adiposity signal acts in the hypothalamus via suppressing food intake and inducing hypermetabolism. Age and obesity are accompanied by leptin resistance. The present study aimed to clarify which components of the catabolic leptin effects are influenced most strongly by aging and which ones by nutritional state-induced alterations in body composition. In our biotelemetric study the effects of a 7-day intracerebroventricular leptin infusion on various parameters of energy balance (food intake, body weight, oxygen consumption, heart rate and body temperature) were analyzed in male Wistar rats of different age-groups (from 3 to 24 months) and nutritional states (normally fed, diet-induced obese and calorie-restricted). Leptin resistance of older animals affected hypermetabolic actions, whereas leptin induced anorexia in all age-groups. Weight reducing effect of leptin diminished in middle-aged and aging animals to become significant again in the oldest group. In diet-induced obese rats leptin-induced hypermetabolism of the young rats and hypermetabolism plus anorexia of the aging ones were suppressed. Calorie-restriction reduced body weight and fat mass to a similar extent in all age-groups. It strongly enhanced leptin-induced hypermetabolism at all ages and prevented the manifestation of anorexigenic actions of leptin with the exception of the oldest group. This latter finding suggests an unexpected increase of responsiveness to anorexigenic leptin actions in old rats. Accordingly, anorexia and hypermetabolism change in disparate ways with aging. Nutritional state predominantly influences hypermetabolic leptin actions. Resistance to both hypermetabolic and anorexigenic actions were promoted by obesity, while calorie-restriction enhanced responsiveness to leptin, especially in old rats.

Hypophysectomy attenuates leptin‐induced tachycardia without affecting leptin's action on appetite and body weight.

Leptin, a hormone produced by adipocytes, regulates energy homeostasis and cardiovascular function by activating its receptors in the central nervous system. Since leptin also modulates pituitary function we examined the role of pituitary hormones in contributing to the effects of leptin on energy homeostasis and cardiovascular function. Male 12‐week‐old hypophysectomized Sprague‐Dawley rats (Hypo, n=5) were instrumented with telemetry probes for determination of mean arterial pressure (MAP) and heart rate (HR) 24‐hrs/day and an intracerebroventricular (ICV) cannula was placed in the brain lateral ventricle for continuous infusion of leptin (15 μg/day). Compared to control rats (Ctrl, n=5), Hypo rats were lighter (215±4 vs 415±6 g), had lower MAP and HR (83±4 and 317±2 vs 105±4 mmHg and 339±4 bpm), with similar caloric intake per gram of body weight (174±19 vs 195±10 kcal/kg) and plasma glucose levels (84±4 vs 80±4 mg/dl). Chronic ICV leptin infusion for 7 days reduced caloric intake (−60%) and body weight (−10%) in both groups. Leptin treatment markedly increased HR in Ctrl rats (~25 bpm) while causing only a modest 8 bpm increase in Hypo rats. Leptin also induced a small increase in MAP in both groups (3 mmHg). These results suggest that hypophysectomy attenuates leptin's effect on HR regulation without altering leptin's ability to suppress appetite and to promote weight loss. (NHLBI‐PO1HL51971/ AHA SDG5680016)

Ganglionic blockade does not impair the chronic CNS‐mediated antidiabetic action of leptin in streptozotocin‐induced diabetic rats

This study examined the role of adrenergic and non‐adrenergic factors released by sympathetic fibers in the CNS‐mediated chronic antidiabetic effects of leptin. Male Sprague‐Dawley rats were instrumented with telemetry probes and arterial and venous catheters for determination of blood pressure (BP), heart rate (HR), blood sampling and intravenous (IV) infusions. An intracerebroventricular (ICV) cannula was placed into the brain lateral ventricle for infusion of leptin or vehicle. After control measurements, STZ was injected IV (50 mg/kg) to induce diabetes and 5 days later leptin (15 μg/d, n=5) or saline vehicle (n=4) were infused ICV for 12 days via osmotic pumps. Beginning on day 6 of treatment, the ganglionic blocker hexamethonium (15 mg/kg/hr) was infused IV for 6 days. Induction of diabetes was associated with hyperglycemia (350±19 mg/dL), hyperphagia (43±3 g/d), bradycardia (−70±11 bpm), polyuria and increased water consumption, which were fully reversed by ICV leptin infusion. Although hexamethonium markedly attenuated leptin's effect on HR, it failed to impair leptin's ability to restore euglycemia (66±2 mg/dL) or to prevent the hyperphagia, polyuria or increased water intake in STZ‐diabetic rats. These results suggest that leptin's powerful CNS antidiabetic actions are mediated via non‐autonomic mechanisms. (NHLBI‐PO1 HL51971, AHA)