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Insulin delivery to the brain has emerged as an important therapeutic target for cognitive disorders associated with abnormal brain energy metabolism. Although insulin is transported across the blood-brain barrier, peripheral routes of administration are problematic due to systemic effects of insulin on blood glucose. Intranasal (IN) administration is being investigated as an alternative route. We conducted a head-to-head comparison of subcutaneous (SC) and IN insulin, assessing plasma and brain pharmacokinetics and blood glucose levels in the mouse. SC insulin (2.4 IU) achieved therapeutically relevant concentrations in the brain (AUC brain = 2537 h·μIU/mL) but dramatically increased plasma insulin (AUC plasma = 520 351 h·*μIU/mL), resulting in severe hypoglycemia and in some cases death. IN administration of the same dose resulted in similar insulin levels in the brain (AUC brain = 3442 h·μIU/mL) but substantially lower plasma concentrations (AUC plasma = 354 h·μIU/mL), amounting to a ∼ 2000-fold increase in the AUC brain:plasma ratio relative to SC. IN dosing also had no significant effect on blood glucose. When administered daily for 9 days, IN insulin increased brain glucose and energy metabolite concentrations (e.g., adenosine triphosphate and phosphocreatine) without causing overt toxicity, suggesting that IN insulin may be a safe therapeutic option for cognitively impaired patients.

Insulin detemir (DET) reduces glycemia comparably to other long-acting insulin formulations but causes less weight gain. Insulin signaling in the brain is catabolic, reducing food intake. We hypothesized that DET reduces weight gain, relative to other insulins, owing to increased transport into the central nervous system and/or increased catabolic action within the brain. Transport of DET and NPH insulin into the cerebrospinal fluid (CSF) was compared over several hours and after the administration of different doses peripherally in rats. DET and NPH had comparable saturable, receptor-mediated transport into the CSF. CSF insulin remained elevated significantly longer after intraperitoneal DET than after NPH. When administered acutely into the 3rd cerebral ventricle, both DET and NPH insulin reduced food intake and body weight at 24 h, and both food intake and body weight remained lower after DET than after NPH after 48 h. In direct comparison with another long-acting insulin, insulin glargine (GLAR), DET led to more prolonged increases in CSF insulin despite a shorter plasma half-life in both rats and mice. Additionally, peripheral DET administration reduced weight gain and increased CSF insulin compared with saline or GLAR in mice. Overall, these data support the hypothesis that DET has distinct effects on energy balance through enhanced and prolonged centrally mediated reduction of food intake., (© 2015 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.)

Objectives: It is a matter of debate whether impaired insulin action originates from a defect at the neural level or impaired transport of the hormone into the brain. In this study, we aimed to investigate the effect of aging on insulin concentrations in the periphery and the central nervous system as well as its impact on insulin-dependent brain activity., Methods: Insulin, glucose and albumin concentrations were determined in 160 paired human serum and cerebrospinal fluid (CSF) samples. Additionally, insulin was applied in young and aged mice by subcutaneous injection or intracerebroventricularly to circumvent the blood-brain barrier. Insulin action and cortical activity were assessed by Western blotting and electrocorticography radiotelemetric measurements., Results: In humans, CSF glucose and insulin concentrations were tightly correlated with the respective serum/plasma concentrations. The CSF/serum ratio for insulin was reduced in older subjects while the CSF/serum ratio for albumin increased with age like for most other proteins. Western blot analysis in murine whole brain lysates revealed impaired phosphorylation of AKT (P-AKT) in aged mice following peripheral insulin stimulation whereas P-AKT was comparable to levels in young mice after intracerebroventricular insulin application. As readout for insulin action in the brain, insulin-mediated cortical brain activity instantly increased in young mice subcutaneously injected with insulin but was significantly reduced and delayed in aged mice during the treatment period. When insulin was applied intracerebroventricularly into aged animals, brain activity was readily improved., Conclusions: This study discloses age-dependent changes in insulin CSF/serum ratios in humans. In the elderly, cerebral insulin resistance might be partially attributed to an impaired transport of insulin into the central nervous system.

The pancreatic hormone insulin plays a well-described role in the periphery, based principally on its ability to lower circulating glucose levels via activation of glucose transporters. However, insulin also acts within the central nervous system (CNS) to alter a number of physiological outcomes ranging from energy balance and glucose homeostasis to cognitive performance. Insulin is transported into the CNS by a saturable receptor-mediated process that is proposed to be dependent on the insulin receptor. Transport of insulin into the brain is dependent on numerous factors including diet, glycemia, a diabetic state and notably, obesity. Obesity leads to a marked decrease in insulin transport from the periphery into the CNS and the biological basis of this reduction of transport remains unresolved. Despite decades of research into the effects of central insulin on a wide range of physiological functions and its transport from the periphery to the CNS, numerous questions remain unanswered including which receptor is responsible for transport and the precise mechanisms of action of insulin within the brain., (© 2015 Elsevier Inc. All rights reserved.)

Research in animals and humans has associated Alzheimer's disease (AD) with decreased cerebrospinal fluid levels of insulin in combination with decreased insulin sensitivity (insulin resistance) in the brain. This phenomenon is accompanied by attenuated receptor expression of insulin and insulin-like growth factor, enhanced serine phosphorylation of insulin receptor substrate-1, and impaired transport of insulin across the blood-brain barrier. Moreover, clinical trials have demonstrated that intranasal insulin improves both memory performance and metabolic integrity of the brain in patients suffering from AD or its prodrome, mild cognitive impairment. These results, in conjunction with the finding that insulin mitigates hippocampal synapse vulnerability to beta amyloid, a peptide thought to be causative in the development of AD, provide a strong rationale for hypothesizing that pharmacological strategies bolstering brain insulin signaling, such as intranasal administration of insulin, could have significant potential in the treatment and prevention of AD. With this view in mind, the review at hand will present molecular mechanisms potentially underlying the memory-enhancing and neuroprotective effects of intranasal insulin. Then, we will discuss the results of intranasal insulin studies that have demonstrated that enhancing brain insulin signaling improves memory and learning processes in both cognitively healthy and impaired humans. Finally, we will provide an overview of neuroimaging studies indicating that disturbances in insulin metabolism--such as insulin resistance in obesity, type 2 diabetes and AD--and altered brain responses to insulin are linked to decreased cerebral volume and especially to hippocampal atrophy.

Investigate possible associations of white matter hyperintensities (WMHs) with the metabolism of cholesterol and insulin in two subgroups of patients with memory complaints and different CSF Aβ42 and CSF tau levels. 59 patients from the memory clinic at Karolinska Hospital were included. Degree of WMHs was rated using the ARWMC scale and the following biomarkers were measured in CSF and plasma: insulin, cholesterol, lanosterol, lathosterol, and oxidized cholesterol metabolites. The WMHs in CSF control-like group correlated with increased brain cholesterol synthesis and reduced efflux of oxysterols and insulin in CSF. In the CSF AD-like group, the WMHs correlated with increased peripheral cholesterol metabolism. Despite having similar appearance on FLAIR images, the pathogenic mechanisms of WMHS are likely to be different in the two groups investigated., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Pregnancy impairs baroreflex gain, but the mechanism is incompletely understood. To test the hypothesis that reductions in brain insulin contribute, we determined whether pregnant rats exhibit lower cerebrospinal fluid (CSF) insulin concentrations and whether intracerebroventricular infusion of insulin normalizes gain of baroreflex control of heart rate in conscious pregnant rats. CSF insulin was lower in pregnant (68 ± 21 pg/mL) compared to virgin (169 ± 25 pg/mL) rats (P < 0.05). Pregnancy reduced baroreflex gain (pregnant 2.4 ± 0.2 bpm/mm Hg, virgin 4.6 ± 0.3 bpm/mm Hg; P < 0.0001) and the maximum heart rate elicited by hypotension (pregnant 455 ± 15 bpm, virgin 507 ± 12 bpm; P = 0.01). Infusion of insulin (100 μU/min) intracerebroventricularly increased baroreflex gain in pregnant (2.4 ± 0.4 to 3.9 ± 0.5 bpm/mm Hg; P < 0.01) but not virgin (4.6 ± 0.4 to 4.2 ± 0.4 bpm/mm Hg; NS) rats. Maximum heart rate was not altered by intracerebroventricular insulin in either group. Interestingly, while in pregnant rats the baroreflex was unchanged by intracerebroventricular infusion of the artificial CSF vehicle, in virgin rats, vehicle infusion lowered baroreflex gain (4.7 ± 0.3 to 3.9 ± 0.3 bpm/mm Hg; P < 0.05) and the maximum baroreflex heart rate (495 ± 19 to 444 ± 21 bpm; P < 0.05). These data support the hypothesis that brain insulin is required to support optimal baroreflex function and that a decrease in brain insulin contributes to the fall in baroreflex gain during pregnancy.

Insulin transported from plasma into the central nervous system (CNS) is hypothesized to contribute to the negative feedback regulation of body adiposity. Because CNS insulin uptake is likely mediated by insulin receptors, physiological interventions that impair insulin action in the periphery might also reduce the efficiency of CNS insulin uptake and predispose to weight gain. We hypothesized that high-fat feeding, which both reduces insulin sensitivity in peripheral tissues and favors weight gain, reduces the efficiency of insulin uptake from plasma into the CNS. To test this hypothesis, we estimated parameters for cerebrospinal fluid (CSF) insulin uptake and clearance during an intravenous insulin infusion using compartmental modeling in 10 dogs before and after 7 weeks of high-fat feeding. These parameters, together with 24-h plasma insulin levels measured during ad libitum feeding, also permitted estimates of relative CNS insulin concentrations. The percent changes of adiposity, body weight, and food intake after high-fat feeding were each inversely associated with the percent changes of the parameter k1k2, which reflects the efficiency of CNS insulin uptake from plasma (r = -0.74, -0.69, -0.63; P = 0.015, 0.03, and 0.05, respectively). These findings were supported by a non-model-based calculation of CNS insulin uptake: the CSF-to-plasma insulin ratio during the insulin infusion. This ratio changed in association with changes of k1k2 (r = 0.84, P = 0.002), body weight (r = -0.66, P = 0.04), and relative adiposity (r = -0.72, P = 0.02). By comparison, changes in insulin sensitivity, according to minimal model analysis, were not associated with changes in k1k2, suggesting that these parameters are not regulated in parallel. During high-fat feeding, there was a 60% reduction of the estimated CNS insulin level (P = 0.04), and this estimate was inversely associated with percent changes in body weight (r = -0.71, P = 0.03). These results demonstrate that increased food intake and weight gain during high-fat feeding are associated with and may be causally related to reduced insulin delivery into the CNS.

Insulin is believed to act as a central adiposity signal by binding to hypothalamic and other brain insulin receptors. Entry of circulating insulin into the brain is accomplished by a saturable receptor-mediated transendothelial transport system and is believed to be impaired in hyperinsulinemic, insulin-resistant, and hyperphagic obese Zucker rats. Theoretically, if hyperinsulinemia is decreased simultaneously while brain capillary insulin binding is increased, uptake of insulin into the brain can be enhanced leading to reduced food intake. To test this hypothesis, we administered diazoxide (DZ, 150 mg/kg/day), an inhibitor of glucose-mediated insulin secretion, or vehicle (control) to 7-week-old female obese and lean Zucker rats for 4 weeks (n = 24-28/subgroup-strain). Animals were assigned to either fasted (FD) or free-fed (FF) protocol for determination of plasma and cerebrospinal fluid (CSF) insulin and brain capillary insulin binding at the end of 4 weeks. DZ obese consumed fewer calories (P<0.01) and gained less weight than control obese (P<0.01), whereas DZ lean had similar amounts of caloric intake and weight gain compared with lean controls. DZ obese had lower fasting and random plasma glucose than control obese (P<0.05). FD and FF DZ-treated obese and lean rats had lower plasma insulin than their respective obese (P<0.01) and lean (P<0.01) controls. FD and FF DZ-treated obese rats demonstrated higher CSF insulin (P<0.05) and CSF/ plasma insulin ratio (P<0.01) than their controls, while only FF DZ lean animals showed higher CSF/plasma insulin ratio (P<0.01) than their controls (P<0.05). This was associated with enhanced brain capillary insulin binding in FD and FF DZ-treated obese (P<0.01) and lean (P<0.05) animals compared with their respective controls. It was concluded that DZ treatment in obese Zucker rats caused a decrease in insulin secretion and partially reversed impaired insulin binding to brain capillaries, leading to enhanced brain insulin uptake, and resulted in reduced food intake and weight gain observed in these animals.

We have previously shown that transport of plasma insulin into the central nervous system (CNS) is mediated by a saturable mechanism consistent with insulin binding to blood-brain barrier insulin receptors and subsequent transcytosis through microvessel endothelial cells. Since glucocorticoids antagonize insulin receptor-mediated actions both peripherally and in the CNS, we hypothesized that glucocorticoids also impair CNS insulin transport. Nine dogs were studied both in the control condition and after 7 days of high-dose oral dexamethasone (DEX) administration (12 mg/day) by obtaining plasma and cerebrospinal fluid (CSF) samples over 8 h for determination of immunoreactive insulin levels during a 90-min euglycemic intravenous insulin infusion (plasma insulin approximately 700 pmol/l). From these data, the kinetics of CNS insulin uptake and removal were determined using a mathematical model with three components (plasma-->intermediate compartment, hypothesized to be brain interstitial fluid-->CSF). DEX increased basal insulin levels 75% from 24 +/- 6 to 42 +/- 30 pmol/l (P < 0.005) and slightly increased basal glucose levels from 5.0 +/- 0.7 to 5.3 +/- 1.0 mmol/l (P < 0.05). DEX also lowered the model rate constant characterizing CNS insulin transport by 49% from 5.3 x 10(-6) +/- 4.0 x 10(-6) to 2.7 x 10(-6) +/- 1.2 x 10(-6) min-2 (P < or = 0.001). As glucocorticoids are known to reduce CSF turnover, we also hypothesized that the model rate constant associated with CSF insulin removal would be decreased by DEX. As expected, the model rate constant for CSF insulin removal decreased 47% from 0.038 +/- 0.013 to 0.020 +/- 0.088 min-1 (P < or = 0.0005) during DEX treatment. We conclude that DEX impairs CNS insulin transport. This finding supports our hypothesis that insulin receptors participate in the CNS insulin transport process and that this process may be subject to regulation. Moreover, since increasing brain insulin transport reduces food intake and body adiposity, this observation provides a potential mechanism by which glucocorticoid excess leads to increased body adiposity.

By acting in the central nervous system, circulating insulin may regulate food intake and body weight. We have previously shown that the kinetics of insulin uptake from plasma into cerebrospinal fluid (CSF) can best be explained by passage through an intermediate compartment. To determine if transport kinetics into this compartment were consistent with an insulin receptor-mediated transport process, we subjected overnight fasted, anesthetized dogs to euglycemic intravenous insulin infusions for 90 min over a wide range of plasma insulin levels (69-5,064 microU/ml) (n = 10). Plasma and CSF samples were collected over 8 h for determination of immunoreactive insulin levels, and the kinetics of insulin uptake from plasma into CSF were analyzed using a compartmental model with three components (plasma-->intermediate compartment-->CSF). By sampling frequently during rapid changes of plasma and CSF insulin levels, we were able to precisely estimate three parameters (average standard deviation 14%) characterizing the uptake of insulin from plasma, through the intermediate compartment and into CSF (k1k2); insulin entry into CSF and insulin clearance from the intermediate compartment (k2 + k3); and insulin clearance from CSF (k4). At physiologic plasma insulin levels (80 +/- 7.4 microU/ml), k1k2 was determined to be 10.7 x 10(-6) +/- 1.3 x 10(-6) min-2. With increasing plasma levels, however, k1k2 decreased progressively, being reduced sevenfold at supraphysiologic levels (5,064 microU/ml). The apparent KM of this saturation curve was 742 microU/ml (approximately 5 nM). In contrast, the rate constants for insulin removal from the intermediate compartment and from CSF did not vary with plasma insulin (k2 + k3 = 0.011 +/- 0.0019 min-1 and k4 = 0.046 +/- 0.021 min-1). We conclude that delivery of plasma insulin into the central nervous system is saturable, and is likely facilitated by an insulin-receptor mediated transport process.

We delineated the ontogeny of rabbit brain insulin concentrations to define the regulatory role of development on this hormone in the central nervous system. Employing a sensitive ELISA, we observed higher concentrations in the late gestation fetal brain (approximately 80-90 ng/g) and early neonatal brain (approximately 195 ng/g) in comparison to the adult (approximately 32 ng/g; P less than 0.01). Further, we characterized this hormone to determine the identity of insulin (or an insulin-like substance) in brain. Employing porcine/bovine or rabbit insulin as standards, we observed that brain insulin mimicked authentic insulin in its migration on SDS-polyacrylamide and native gel electrophoresis, immunogenicity on Western blot analysis, and its elution profile on immunoaffinity column chromatographic, and high performance liquid chromatographic separation. We then examined the developmental effects on circulating and cerebrospinal fluid (CSF) radioimmunoassayable insulin levels. No statistically significant differences (ANOVA) existed through development in either the serum or CSF insulin levels. Employing multiple regression analysis, no correlation was evident between brain and either serum or CSF insulin concentration. A search for insulin mRNA by Northern blot analysis yielded minute amounts of atypical large sized transcripts. We conclude that the insulin peptide in the central nervous system closely resembles (or is identical to) circulating insulin in many properties and that there is a developmental increase in brain insulin concentrations, the maximal peak occuring in the late gestation fetus and early neonate. Insulin concentrations in brain demonstrate no conventional relationship to either the serum or CSF insulin levels, suggesting an additional source of peptide, which contributes beyond that which is available via the circulation. The amounts of insulin present within the central nervous system are minute (difficult to detect) but in the range (10-100 ng) where the hormone can interact with either insulin or insulin-like growth factor I (IGF-I) receptors that are abundantly present on developing brain cells, thereby executing the biological function of the hormone.

We have provided strong support for the hypothesis that the pancreatic hormone, insulin, provides a signal to the brain indicating the level of adiposity. Because insulin is found in the cerebrospinal fluid (CSF) in direct proportion to plasma levels, and because changes of plasma insulin result in subsequent changes of CSF insulin, we previously hypothesized that the blood-borne insulin signal enters the central nervous system by initially entering the CSF and then diffuses into the brain. Such a route explained the time lag for influences of insulin upon food intake and body weight. Recent evidence suggests that insulin may enter the brain directly through brain capillaries, raising the possibility that what is measured in the CSF may not be indicative of insulin on its way into critical brain areas. Implications of this change of route of entry of insulin into the brain for the regulation of food intake and body weight are discussed.

Injection of insulin to the CSF in the presence of spontaneous diuresis and hydration gives rise to the growth of reabsorption of osmotically free water accompanied by high tubular transport, as well as to the development of antinatriuresis and inhibition of diuresis. Experiments with a preliminary blockade of beta-adrenoreceptors with propranolol or administration of a beta-blocker following insulin injection demonstrated beta-adrenoreactive brain structures to be involved in the mechanism of action of insulin. Secondary activation of vasopressin secretion and release in the blood may be mediated via these structures, as a result of which reabsorption of osmotically free water in renal tubules gets increased. Thus, CSF insulin effects its influence on renal function via the central neurohumoral mechanisms which work due to beta-adrenergic brain receptors.

The present study was performed to explore the role of exogenous insulin in CSF in the control of energy balance in the rat. For this purpose, adult male Sprague-Dawley rats carrying an indwelling cannula in the right lateral cerebral ventricle were infused for a maximum of 10 days with insulin (Actrapid) at various rates (starting at 0, 45, 85, 170, and 600 ng/day) or anti-insulin antibody (IgG fraction; diluted 1:10 wt/vol) with an osmotic minipump. All those treatments did not modify the growing rates; neither total daily food intake nor the circadian rhythm of food intake was further modified. The chronic insulin infusion starting at 600 ng/day resulted in a chronic significant increase in CSF insulin levels without changing the plasma insulin level. It failed to alter specific insulin binding sites to Triton X-100 solubilized microsomal membranes from various brain areas (cerebral cortex, olfactory bulbs, and lateral and medial hypothalami) at the end of the 5- or 10-day period of insulin infusion. Purification of insulin receptors on a wheat germ agglutinin did not reveal any further effect of insulin. From these results, it seems unlikely that the input to the brain insulin-effector systems could arise from CSF insulin.

This article describes the close relationship among the hormone insulin, the central nervous system, and the regulation of food intake and body adiposity. The initial section documents the control of insulin output from the pancreas by the central nervous system, and a later section describes the relationship of insulin levels in the blood to the degree of adiposity. Another section documents the ability of insulin to gain access to the brain and to elicit responses there. Finally, the behavioral effects of insulin added to the brain, and especially its ability to reduce food intake and body weight, is discussed. The implications to obesity are stressed throughout.