The energy metabolism project aims to understand how energy intake and expenditure, and diabetes, affect neuronal plasticity and vulnerability to disease.   These studies include both basic science and translational research components, are integrated with the other 3 CMNS projects, and include collaborations with several LNS and NIA IRP investigators.
 
Diabetes and excessive energy intake adversely affect multiple components of hippocampal plasticity.   We found that, in both insulin-deficient rats and insulin-resistant mice, diabetes impairs hippocampus-dependent memory, perforant path synaptic plasticity and adult neurogenesis, and the adrenal steroid corticosterone contributes to these adverse effects of diabetes (56). Rats treated with streptozocin have reduced insulin and show hyperglycemia, increased corticosterone, and impairments in hippocampal neurogenesis, synaptic plasticity and learning. Similar deficits are observed in db/db mice, which are characterized by insulin resistance, elevated corticosterone and obesity. Changes in hippocampal plasticity and function in both models are reversed when normal physiological levels of corticosterone are maintained, suggesting that cognitive impairment in diabetes may result from glucocorticoid-mediated deficits in neurogenesis and synaptic plasticity.
   We evaluated the performance of young diabetic rats in the 14-unit T-maze, a task that is sensitive to hippocampal deficits (57). To assess the contribution of diabetes-induced elevations in corticosterone levels, we examined maze learning in diabetic rats that had levels of corticosterone 'clamped' through adrenalectomy and low-dose corticosterone replacement. For comparison, we also tested a separate group of young and aged rats in the maze. Adrenally intact diabetic rats learned poorly in the 14-unit T-maze. Preventing the increases in corticosterone levels that accompanies the onset of experimental diabetes also prevented deficits in complex maze learning. The pattern of errors made by adrenally intact diabetic rats was similar to the pattern of errors made by aged rats, suggesting that the cognitive profiles of diabetic and aged rats share common features.
  Rats fed with a high-fat, high-glucose diet supplemented with high-fructose corn syrup showed alterations in energy and lipid metabolism similar to clinical diabetes, with elevated fasting glucose and increased cholesterol and triglycerides. Rats maintained on this diet for 8 months exhibited impaired spatial learning ability, reduced hippocampal dendritic spine density, and reduced LTP at Schaffer collateral--CA1 synapses (58). These changes occurred concurrently with reductions in levels of BDNF in the hippocampus. Thus, a high-calorie diet reduces hippocampal synaptic plasticity and impairs cognitive function, possibly through BDNF-mediated effects on dendritic spines.
 
Exercise and dietary energy restriction counteract the adverse effects of diabetes on hippocampal plasticity.  To investigate whether manipulations that enhance neurotrophin levels will also restore neuronal structure and function in diabetes, we examined the effects of wheel running and dietary energy restriction on hippocampal neuron morphology and BDNF levels in db/db mice, a model of insulin resistant diabetes (59). Running wheel activity, CR, or the combination of the two treatments increased levels of BDNF in the hippocampus of db/db mice. Enhancement of hippocampal BDNF was accompanied by increases in dendritic spine density on the secondary and tertiary dendrites of dentate granule neurons. These studies suggest that diabetes exerts detrimental effects on hippocampal structure, and that this state can be attenuated by increasing energy expenditure and decreasing energy intake.

Metabolic, neuroendocrine, and cognitive responses to dietary energy restriction and excess are sex-dependent.  Females and males typically play different roles in survival of the species and we therefore reasoned that their brains may respond differently to food scarcity or excess. To elucidate the physiological basis of sex differences in responses to energy intake, we maintained groups of male and female rats for 6 months on diets with usual, reduced calories (20% and 40% CR), ADF, or elevated (high-fat/high-glucose) energy levels and measured multiple physiological variables related to reproduction, energy metabolism, and behavior (39). In response to 40% CR, females became emaciated, ceased cycling, underwent endocrine masculinization, exhibited a heightened stress response, increased their spontaneous activity, improved their learning and memory, and maintained elevated levels of circulating BDNF. In contrast, males on 40% CR maintained a higher body weight than the 40% CR females and did not change their activity levels appreciably. Additionally, there was no significant change in the cognitive ability of the males on the 40% CR diet. Males and females exhibited similar responses of circulating lipids (cholesterols/triglycerides) and energy-regulating hormones (insulin, leptin, adiponectin, ghrelin) to 20% CR and ADF, with the changes being quantitatively greater in males. The high-fat/high-glucose diet had no significant effects on most neurological variables measured, but adversely affected the reproductive cycle in females. Heightened cognition and motor activity, combined with reproductive shutdown, in females may maximize the probability of their survival during periods of energy scarcity and may be an evolutionary basis for the vulnerability of women to anorexia nervosa.

GLP-1 signaling protects neurons against dysfunction and degeneration in animal models of neurodegenerative disease. Glucagon-like peptide-1 (GLP-1) is an endogenous insulinotropic peptide secreted from the gastrointestinal tract in response to food intake. It enhances pancreatic islet b-cell proliferation and glucose-dependent insulin secretion, and lowers blood glucose and food intake in patients with type 2 diabetes. A long-acting GLP-1 receptor (GLP-1R) agonist, exendin-4 (Ex-4), is the first of this new class of anti-hyperglycemia drugs approved to treat diabetes. GLP-1Rs are coupled to the cAMP second messenger pathway and, along with pancreatic cells, are expressed within the nervous system of rodents and humans, where receptor activation elicits neurotrophic actions. We detected GLP-1R mRNA expression in both cultured embryonic primary cerebral cortical and ventral mesencephalic (dopaminergic) neurons (30). These cells are vulnerable to hypoxia- and 6-hydroxydopamine-induced cell death, respectively. We found that GLP-1 and Ex-4 conferred protection in these cells, but not in cells from GLP-1R-deficient mice. Administration of Ex-4 reduced brain damage and improved functional outcome in a transient middle cerebral artery occlusion stroke model. Ex-4 treatment also protected dopaminergic neurons against degeneration, preserved dopamine levels, and improved motor function in the MPTP mouse model of PD. Our findings demonstrate that Ex-4 can protect neurons against metabolic and oxidative insults, and they provide preclinical support for the therapeutic potential for Ex-4 in the treatment of stroke and PD.
Huntington's disease (HD) patients exhibit neuronal dysfunction/degeneration, chorea, and progressive weight loss. Additionally, they suffer from abnormalities in energy metabolism affecting both the brain and periphery.  We treated HD mice with the Ex-4, to test whether euglycemia could be achieved, whether pancreatic dysfunction could be alleviated, and whether the mice showed any neurological benefit (40). Blood glucose and insulin levels and various appetite hormone concentrations were measured during the study. Additionally, motor performance and life span were quantified and mutant huntingtin (mhtt) aggregates were measured in both the pancreas and brain. Ex-4 treatment ameliorated abnormalities in peripheral glucose regulation and suppressed cellular pathology in both brain and pancreas in a mouse model of HD. The treatment also improved motor function and extended the survival time of the HD mice. These clinical improvements were correlated with reduced accumulation of mhtt protein aggregates in both islet and brain cells. Targeting both peripheral and neuronal deficits, Ex-4 is an attractive agent for therapeutic intervention in HD patients.

We also found that Ex-4 treatment can ameliorate several pathological processes in diabetic 3xTgAD mice (31).  When 11 to 12.5 month old female 3xTg AD mice were challenged with streptozotocin or saline, and thereafter treated with a continuous subcutaneous infusion of Ex-4 or vehicle, Ex-4 ameliorated the diabetic effects of streptozotocin in 3xTg-AD mice, elevating plasma insulin and lowering both plasma glucose and hemoglobin A1c (HbA1c) levels. Furthermore, brain levels of Ab precursor protein and Ab, which were elevated in diabetic 3xTg-AD mice, were significantly reduced in Ex-4 treated mice. Together, these results suggest a potential value of Ex-4 in AD, particularly when associated with diabetes or glucose intolerance.

Autonomic function and dysfunction:  a window into the early stages of neurodegenerative disease?  In previous studies we continuously monitored the heart rate,  blood pressure, body temperature and movement of mice and rats maintained on ad libitum, ADF or 40% CR diets.  We found that resting blood pressure and heart rate were gradually reduced, and heart rate variability was increased, during a 2 – 4 week on diet period in animals on the ADF and CR diets compared to the control diet (37, 66).   We further showed that animals on the ADF diet exhibit improved cardiovascular stress adaptation when challenged with either immobilization stress or cold water swim stress (66).    The beneficial effects of ADF and CR diets were lost within 1 – 2 weeks of reversion to the ad libitum control diet.   More recently we have been elucidating ANS abnormalities in mouse models of AD, PD and HD.  HD patients exhibit profound ANS dysfunction including dysregulation of cardiovascular control which often precedes psychiatric or motor symptoms.   We therefore examined central heart rate control in N171-82Q mice, a mouse model of HD (CMNS Appendix D).  Resting heart rate was elevated in HD mice at presymptomatic and early disease stages, while heart rate responses were significantly attenuated in HD mice during restraint stress compared to wild type control mice, suggesting that central control of heart rate is dysfunctional in HD mice. BDNF and TrkB protein levels were significantly decreased in ANS cardiovascular nuclei in the pons and medulla of HD mice across disease stages as well as in human HD medulla.  Central administration of BDNF restored the heart rate to control levels.  These findings suggest a functional link between diminished BDNF expression in brainstem regions responsible for central cardiovascular control and abnormal heart rates in HD mice, and suggest a novel therapeutic target for correcting ANS dysfunction in HD.

A cautionary note for lab animal research: “Control” animals are metabolically morbid. We published a perspective article in PNAS intended to raise awareness of the fact that most standard ‘control’ rats and mice used in biomedical research are sedentary, obese, glucose intolerant, and on a trajectory to premature death (42).  This metabolic state of the animals may in some instances confound data interpretation and outcomes of human studies. Fundamental aspects of cellular physiology, vulnerability to oxidative stress, inflammation, and associated diseases are among the many biological processes affected by dietary energy intake and exercise. Although overfed sedentary rodents may be reasonable models for the study of obesity in humans, treatments shown to be efficacious in these animal models may prove ineffective or exhibit novel side effects in active, normal-weight subjects.  Ideally, many experiments should be designed with two control groups, the standard control and a leaner more fit control group that is fed less and exercises more.