Cerebrospinal Fluid Insulin Levels Research Articles

Association of cigarette smoking with cerebrospinal fluid biomarkers of insulin sensitivity and neurodegeneration.

Cigarette smoking increases both the risk for insulin resistance and amyloid-β (Aβ) aggregation, and impaired brain insulin/insulin-like growth factor 1 (IGF1) signaling might increase risk factors for Alzheimer's disease (AD). We aimed to investigate the association among cerebrospinal fluid (CSF) insulin sensitivity/IGF1, glucose/lactate, and Aβ42 and further explore whether insulin sensitivity contributed to the risk for AD in active smokers. In this cross-sectional study, levels of insulin, IGF1, and lactate/glucose of 75 active smokers and 78 nonsmokers in CSF were measured. Three polymorphisms regulating IGF1 were genotyped. Analysis of variance was used to compare differences of variables between groups. Partial correlation was performed to test the relationship between CSF biomarkers and smoking status. General linear models were applied to test the interaction of the effect of single nucleotide polymorphisms and cigarette smoking on CSF IGF1 levels. In the CSF from active smokers, IGF1 and lactate levels were significantly lower (p=.016 and p=.010, respectively), whereas Aβ42 (derived from our earlier research) and insulin levels were significantly higher (p<.001 and p=.022, respectively) as compared to the CSF from nonsmokers. The AG+GG genotype of rs6218 in active smokers had a significant effect on lower CSF IGF1 levels (p=.004) and lower CSF insulin levels in nonsmokers (p=.016). Cigarette smoking as the "at-risk" factor for AD might be due to lower cerebral insulin sensitivity in CSF, and the subjects with rs6218G allele seem to be more susceptible to the neurodegenerative risks for cigarette smoking.

Growth differentiation factor 15 increases in both cerebrospinal fluid and serum during pregnancy.

AimGrowth differentiation factor 15 (GDF15) increases in serum during pregnancy to levels not seen in any other physiological state and is suggested to be involved in pregnancy-induced nausea, weight regulation and glucose metabolism. The main action of GDF15 is regulated through a receptor of the brainstem, i.e., through exposure of GDF15 in both blood and cerebrospinal fluid (CSF). The aim of the current study was to measure GDF15 in both CSF and serum during pregnancy, and to compare it longitudinally to non-pregnant levels.MethodsWomen were sampled at elective caesarean section (n = 45, BMI = 28.1±5.0) and were followed up 5 years after pregnancy (n = 25). GDF15, insulin and leptin were measured in CSF and serum. Additional measurements included plasma glucose, and serum adiponectin and Hs-CRP.ResultsGDF15 levels were higher during pregnancy compared with follow-up in both CSF (385±128 vs. 115±32 ng/l, P<0.001) and serum (73789±29198 vs. 404±102 ng/l, P<0.001). CSF levels correlated with serum levels during pregnancy (P<0.001), but not in the non-pregnant state (P = 0.98). Both CSF and serum GDF15 were highest in women carrying a female fetus (P<0.001). Serum GDF15 correlated with the homeostatic model assessment for beta-cell function and placental weight, and CSF GDF15 correlated inversely with CSF insulin levels.ConclusionThis, the first study to measure CSF GDF15 during pregnancy, demonstrated increased GDF15 levels in both serum and CSF during pregnancy. The results suggest that effects of GDF15 during pregnancy can be mediated by increases in both CSF and serum levels.

1719-P: Novel Mass Spectrometry–Based Selected Reaction Monitoring Proteomics for Analysis of Low Abundant Insulin Levels in Cerebrospinal Fluid

Circulating insulin levels are routinely measured using Enzyme-linked Immunosorbent Assay (ELISA). However, ELISA is not sensitive enough to measure cerebrospinal fluid (CSF) insulin levels which are 10-100 times lower than those of plasma: yet ELISA results are often reported for insulin analysis in CSF where it serves as a surrogate for brain interstitial fluid insulin levels. We developed a novel LC-MS/MS based selected reaction monitoring (SRM) proteomics assay for quantitative analyses targeting predetermined peptides derived from trypsin digests of insulin; each peptide depends on specific SRM transitions. We determined insulin levels using our novel insulin SRM proteomics method in the matching plasma and CSF samples that were from individuals fasting ≥12 hours and individuals after 2 hours of steady state hyperglycemia by clamp technology (2 hr SS HG). All samples were obtained in individuals with normal fasting glucose and HbA1C levels. Statistically significant differences were observed in both plasma and CSF between fasting and 2 hr SS HG by two tailed and unpaired student t test and 2-way matched ANOVA as shown in Table below. Our results confirmed that peripheral endogenous insulin appears in CSF at least 2 hours after steady state hyperglycemia in subjects with normal glucose tolerance. Our SRM insulin tryptic peptide panel reliably quantified insulin peptide levels in human CSF samples. Disclosure Q. Liu: None. M. Zhu: None. C.W. Chia: None. J.F. O’Connell: None. P. Zhang: None. J.M. Egan: None. Funding National Institute on Aging

Characterizing the Effects of Diabetes and Obesity on Insulin and Leptin Levels amongst Pregnant Women.

In this study, we assess the impact of obesity and diabetes on maternal brain and periphery, as well as fetal exposure to insulin and leptin, and two hormones that play an important role in regulating energy homeostasis. Fasting maternal plasma, fetal cord vein and artery plasma, and maternal cerebrospinal fluid (CSF) were collected in 37 women (12 lean, nondiabetic [prepregnancy body mass index (BMI): 22.9 ± 1.7 kg/m2]; 12 overweight/obese nondiabetic [BMI: 37.8 ± 7.3 kg/m2]; 13 gestational/type 2 diabetes mellitus [BMI: 29.8 ± 7.3 kg/m2]) with uncomplicated singleton pregnancies undergoing elective Cesarean delivery. HbA1C, insulin, glucose, and leptin levels were measured. Compared with lean mothers, mothers with obesity and diabetes mellitus (DM) had significantly lower CSF-to-plasma ratios of insulin. Moreover, mothers with obesity and DM had significantly lower cord arterial and cord venous to maternal plasma ratios of insulin, but not leptin, compared with lean mothers. There were no differences in CSF and cord blood insulin and leptin levels between obese and DM mothers. Compared with lean individuals, mothers with obesity and DM have relative deficiencies in insulin exposure. The patterns observed in mothers with obesity and diabetes were similar highlighting the importance of the maternal metabolic environment in obesity and suggesting obese patients warrant further clinical focus.

Resistance to the sympathoexcitatory effects of insulin and leptin in late pregnant rats.

Pregnancy increases sympathetic nerve activity (SNA), although the mechanisms responsible for this remain unknown. We tested whether insulin or leptin, two sympathoexcitatory hormones increased during pregnancy, contribute to this. Transport of insulin across the blood-brain barrier in some brain regions, and into the cerebrospinal fluid (CSF), was increased, although brain insulin degradation was also increased. As a result, brain and CSF insulin levels were not different between pregnant and non-pregnant rats. The sympathoexcitatory responses to insulin and leptin were abolished in pregnant rats. Blockade of arcuate nucleus insulin receptors did not lower SNA in pregnant or non-pregnant rats. Collectively, these data suggest that pregnancy renders the brain resistant to the sympathoexcitatory effects of insulin and leptin, and that these hormones do not mediate pregnancy-induced sympathoexcitation. Increased muscle SNA stimulates glucose uptake. Therefore, during pregnancy, peripheral insulin resistance coupled with blunted insulin- and leptin-induced sympathoexcitation ensures adequate delivery of glucose to the fetus. Pregnancy increases basal sympathetic nerve activity (SNA), although the mechanism responsible for this remains unknown. Insulin and leptin are two sympathoexcitatory hormones that increase during pregnancy, yet, pregnancy impairs central insulin- and leptin-induced signalling. Therefore, to test whether insulin or leptin contribute to basal sympathoexcitation or, instead, whether pregnancy induces resistance to the sympathoexcitatory effects of insulin and leptin, we investigated α-chloralose anaesthetized late pregnant rats, which exhibited increases in lumbar SNA (LSNA), splanchnic SNA and heart rate (HR) compared to non-pregnant animals. In pregnant rats, transport of insulin into cerebrospinal fluid and across the blood-brain barrier in some brain regions increased, although brain insulin degradation was also increased; brain and cerebrospinal fluid insulin levels were not different between pregnant and non-pregnant rats. Although i.c.v. insulin increased LSNA and HR and baroreflex control of LSNA and HR in non-pregnant rats, these effects were abolished in pregnant rats. In parallel, pregnancy completely prevented the actions of leptin with respect to increasing lumbar, splanchnic and renal SNA, as well as baroreflex control of SNA. Blockade of insulin receptors (with S961) in the arcuate nucleus, the site of action of insulin, did not decrease LSNA in pregnant rats, despite blocking the effects of exogenous insulin. Thus, pregnancy is associated with central resistance to insulin and leptin, and these hormones are not responsible for the increased basal SNA of pregnancy. Because increases in LSNA to skeletal muscle stimulates glucose uptake, blunted insulin- and leptin-induced sympathoexcitation reinforces systemic insulin resistance, thereby increasing the delivery of glucose to the fetus.

Pioglitazone ameliorates Aβ42 deposition in rats with diet-induced insulin resistance associated with AKT/GSK3β activation.

Pioglitazone may have potential benefits as an alternative therapeutic treatment for patients with Alzheimer's disease (AD), particularly in individuals that also have comorbid diabetes; however, the mechanisms of action remain unclear. The present study aimed to explore the effects of pioglitazone on amyloid β, isoform 42 (Aβ42) deposition in rats with diet-induced insulin resistance (IR). Diet-induced IR model rats were established in the presence or absence of pioglitazone. Plasma glucose and insulin levels, and cerebrospinal fluid insulin levels were measured; in addition, hippocampal tissues were collected for immunohistochemical analysis of Aβ42 expression. The levels of insulin-degrading enzyme (IDE) and peroxisome proliferator-activated receptor γ (PPARγ) mRNA and protein expression were analyzed by reverse transcription-quantitative polymerase chain reaction and western blotting, respectively. In addition, the activation of glycogen synthase kinase 3β (GSK3β) induced by phosphatidylinositol 3-kinase (PI3K) /protein kinase B (AKT) signaling was detected by western blotting. Results from the present study demonstrated that pioglitazone may enhance peripheral and brain insulin sensitivity in diet-induced IR model rats. Treatment with pioglitazone ameliorated Aβ42 deposition in the hippocampus by increasing IDE and PPARγ expression. Notably, activation of the PI3K/AKT/GSK3β pathway was also demonstrated to serve a role in pioglitazone-induced Aβ42 degradation, which was abrogated by the PPARγ antagonist GW9662. Results from the present study indicated that pioglitazone may improve insulin sensitivity and ameliorate Aβ42 accumulation in rats with diet-induced IR by regulating AKT/GSK3β activation, suggesting that pioglitazone may be a promising drug for AD treatment.

Relationship between Insulin Levels and Nonpsychotic Dementia: A Systematic Review and Meta-Analysis

Objectives To explore the relationship between insulin levels and nonpsychotic dementia. Methods Six electronic databases (PubMed, Cochrane, SCI, CNKI, VIP, and Wanfang) were searched from January 1, 2007, to March 1, 2017. Experimental or observational studies that enrolled people with nonpsychotic dementia or abnormal insulin levels in which insulin levels or MMSE scores (events in nonpsychotic dementia) were the outcome measures. Random-effects models were chosen for this meta-analysis. Sample size, mean, s.d., and events were primarily used to generate effect sizes (with the PRIMA registration number CRD42017069860). Results 50 articles met the final inclusion criteria. Insulin levels in cerebrospinal fluid were lower (Hedges' g = 1.196, 95% CI = 0.238 to 2.514, and P = 0.014), while the levels in peripheral blood were higher in nonpsychotic dementia patients (Hedges' g = 0.853 and 95% CI = 0.579 to 1.127), and MMSE scores were significantly lower in the high insulin group than in the healthy control group (Hedges' g = 0.334, 95% CI = 0.249 to 0.419, and P = 0.000). Conclusions Our comprehensive results indicate that blood insulin levels may increase in patients with nonpsychotic dementia.

Cerebrospinal fluid levels of insulin, leptin, and agouti-related protein in relation to BMI in pregnant women.

During pregnancy, metabolic interactions must be adapted, though neuroendocrine mechanisms for increased food intake are poorly understood. The objective of this study was to characterize differences in insulin, leptin, and agouti-related protein (AgRP) levels in serum and cerebrospinal fluid (CSF) in pregnant women with normal weight (NW) and pregnant women with overweight (OW) or obesity (OB). Placenta as a source for increased peripheral AgRP levels during pregnancy was also investigated. Women were recruited at admission for elective cesarean section. Insulin, AgRP, and leptin were measured in serum and CSF from 30 NW, 25 OW, and 21 OB at term. Serum during pregnancy and placenta at term were collected for further AgRP analysis. Immunohistology showed placental production of AgRP and serum AgRP levels increased throughout pregnancy. CSF AgRP, leptin, and insulin levels were higher in OW and OB than NW. Serum leptin and insulin levels were higher and AgRP lower in OB than NW. High serum AgRP levels might protect from the suppressive effects of leptin during pregnancy. Pregnant women with OB and OW might further be protected from the suppressive effect of leptin by high CSF AgRP levels. Evidence was found, for the first time, of human placental AgRP production mirrored by levels in the circulation.

Novel CSF Biomarker of Metabolic Dysfunction Predicts AD‐like Associations across the Alzheimer's Spectrum

Obesity and insulin resistance(IR) are associated with brain atrophy and cognitive decline, particularly in Alzheimer's disease (AD). IR is correlated with temporal and frontal amyloid deposition and brain atrophy in late middle‐aged participants at risk for AD, as well as less glucose metabolism in cognitively normal elders and AD participants. Because insulin‐degrading enzyme complicates the measurement of insulin levels in cerebrospinal fluid (CSF), an alternative biomarker for metabolic dysregulation in CSF should be used to draw conclusions between central hyper insulinemia and associations with brain atrophy and hypometabolism. Ecto‐nucleotide pyrophosphatase/phosphodiesterase 2 (ENPP2), also known as autotaxin, is studied in tumor cell mobility and is produced by beige adipose tissue. ENPP2 regulates energy metabolism, and is higher in AD prefrontal cortex (PFC). Additionally, ENPP2 stimulates the formation of lysophosphatidic acid, which promotes the generation of tissue fibrosis in vivo and in vitro. We studied Alzheimer's Disease Neuroimaging Initiative(ADNI) participants who were cognitively normal (CN; n=86) or had Mild Cognitive impairment (MCI; n=135) or AD (n=66). Statistical analyses were conducted using SPSS software. Multinomial regression analyses tested if higher ENPP2 was associated with higher relative risk ratios for MCI or AD diagnosis, relative to the CN reference group. Mixed model analyses were used to regress ENPP2 against metabolic, MRI, FDG‐PET and cognitive outcomes. Then on‐parametric Spearman's statistic was used to correlate ENPP2 and CSF biomarker values. T1‐weighted MRI images were preprocessed using FreeSurfer 4.3, and cortical thickness was examined in8 bilateral regions of interest. CSF ENPP2 levels strongly corresponded with CSF insulin‐like growth factor binding protein 2, an established biomarker of glucoregulatory function, and systemic fasting glucose. ENPP2 levels were higher in MCI and AD, and each point increase in log‐based ENPP2 values corresponded to a 3.5 to 5 times increase in the odds of having some degree of clinically relevant memory impairment. Higher ENPP2 predicted thinner PFC cortical thickness in most regions of interest in AD such as medialorbitofrontal cortex, and pars orbital is in MCI. In addition, less bilateral prefrontal cortex glucose metabolism was seen in AD. ENPP2 was also positively correlated with levels of total tau, p‐Tau181, total tau/Aβ1‐42, as well as pTau‐181/Aβ1‐42 to a marginal degree. This pattern suggests that CSF ENPP2 predicts AD neuropathology in a manner similar to dysmetabolism. Future studies should examine the potential benefit of decreasing ENPP2 levels in individuals with MCI or AD.Support or Funding InformationThis study was funded by Iowa State University and NIH K99 AG047282. Neither funding source had any involvement in the report. Data collection and sharing for this project were funded by the ADNI (National Institutes of Health Grant U01‐AG‐024904) and Department of Defense ADNI (award number W81XWH‐12‐2‐0012). ADNI is funded by the National Institute on Aging, the National Institute of Biomedical Imaging and Bioengineering, and through generous contributions from the Alzheimer's Association and the Alzheimer's Drug Discovery Foundation. The Canadian Institutes of Health Research is providing funds to support ADNI clinical sites in Canada. Private‐sector contributions are facilitated by the Foundation for the National Institutes of Health (www.fnih.org). The grantee organization is the Northern California Institute for Research and Education, and the study is coordinated by the Alzheimer's Disease Cooperative Study at the University of California, San Diego. ADNI data are disseminated by the Laboratory for Neuro Imaging at the University of Southern California. The data used in the preparation of this article were obtained from the ADNI database (adni.loni.usc.edu). As such, the investigators within the ADNI contributed to the design and implementation of ADNI and/or provided data but did not participate in analysis or writing of this report.

Increased insulin-like growth factor-1 levels in cerebrospinal fluid of advanced subacute sclerosing panencephalitis patients.

Subacute sclerosing panencephalitis (SSPE) is a progressive, lethal disease. Brain histopathology in certain SSPE patients shows, neurofibrillary tangles composed of abnormally phosphorylated, microtubule-associated protein tau (PHF-tau). Because the, phosphorylation of tau is inhibited by insulin and insulin-like growth factor-1 (IGF-1), we investigated cerebrospinal fluid (CSF) insulin and IGF-1 levels in SSPE patients. In this study CSF IGF-1 and insulin levels of 45 SSPE and 25 age-matched control patients were investigated. CSF IGF-1 levels were significantly higher in SSPE patients at stage 4, compared to other stages (p 0.05). CSF insulin and IGF-1 levels were both positively correlated with serum measles IgG. The correlation between CSF insulin and IGF-1 levels and serum measles virus IgG titer may be the result of, insulin activating IGF-1 receptors, and consequently, IGF-1 stimulating, plasma cells and enhancing IgG production. Increased IGF-1 may also, inhibit the phosphorylation of tau. Further studies examining the, correlation between IGF-1, insulin, tau, and PHF-tau levels in the same, patients may clarify any possible pathogenetic relation between these, pathways.

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Pioglitazone Ameliorates Intracerebral Insulin Resistance and Tau-protein Hyperphosphorylation in Rats with Type 2 Diabetes

To investigate intracerebral insulin resistance and its relationship with tau-protein hyperphosphorylation. A rat model of type 2 diabetes (T2D) was established with streptozotocin (STZ). Diabetic rats received intragastric administration of pioglitazone (PIO group) or normal saline (T2D group) for 4 weeks. As a control, non-diabetic rats received intragastric normal saline (CTL group). The insulin concentrations in cerebrospinal fluid (CSF) and blood were determined with radioimmunoassay, and blood glucose concentration was determined using a glucose oxidation technique. Total and phosphorylated levels of protein kinase B (AKT), glycogen synthase kinase-3β (GSK-3β) and tau-protein in the hippocampus were analyzed using western blotting. The plasma insulin level in the T2D group was higher, and the CSF insulin level in the T2D group lower than in the CTL group. Hippocampal phosphorylated AKT and phosphorylated GSK-3β levels were significantly lower in the T2D group than in the CTL group. Hippocampal tau-protein in the T2D group was hyperphosphorylated at Ser199 and Ser396. Plasma insulin levels in the PIO group were lower than in the T2D group, with no differences in CSF insulin levels. Phosphorylated AKT and phosphorylated GSK-3β levels in the PIO group were significantly higher than in the T2D group Hippocampal phosphorylated tau-protein (Ser199/Ser396) was lower in the PIO group than in the T2D group. Hyperphosphorylation of tau-protein in pioglitazone-treated rats with T2D was improved. Rats with T2D have both cerebral insulin resistance and cerebral hypoinsulinism. Pioglitazone can ameliorate intracerebral insulin resistance and decrease tau-protein hyperphosphorylation, but cannot increase intracerebral insulin levels.

Insulin Levels are Decreased in the Cerebrospinal Fluid of Women with Prodomal Alzheimer's Disease

Previous studies have failed to reach consensus on insulin levels in cerebrospinal fluid of Alzheimer's disease (AD) patients and on its relation to pathological features. We performed a new analysis in patients at different stages of AD, and investigated the relationship of insulin levels with biochemical disease markers and with cognitive score. We included 99 patients from our Memory Clinic (Karolinska University Hospital, Sweden), including: 27 patients with mild AD, 13 that progressed from mild cognitive impairment (MCI) to AD in two years time, 26 with MCI stable after two years, and 33 with subjective cognitive impairment. Insulin was significantly decreased in the cerebrospinal fluid of both women and men with mild AD. Insulin deficits were seen in women belonging to both MCI groups, suggesting that this occurs earlier than in men. Insulin was positively associated with amyloid-β 1-42 (Aβ1-42) levels and cognitive score. Furthermore, total-tau/(Aβ1-42*insulin) ratio showed strikingly better sensitivity and specificity than the total-tau/Aβ1-42 ratio for early AD diagnosis in women.

Low cerebrospinal fluid insulin levels in obese humans

To the Editor: In healthy men, insulin concentrations in cerebrospinal fluid (CSF) increase after acute elevations of plasma insulin levels, indicating that the brain content of insulin is normally finely tuned to circulating insulin [1]. Central nervous insulin promotes weight loss [2] and attenuates peripheral insulin resistance via inhibition of hepatic gluconeogenesis [3]. With increasing body weight, plasma insulin levels rise as a result of elevated insulin resistance. It is not yet known whether in hyperinsulinaemic obese subjects this increase is accompanied by parallel increases in CSF insulin levels that would act to promote weight loss in these patients. We compared insulin concentrations in plasma and CSF in a group of 45 subjects (23 women, age 19–80 years) with a wide body weight range (BMI 16.24–38.10 kg/m). Five subjects were newly diagnosed as diabetic by elevated fasting plasma glucose levels (>7.0 mmol/l). Since their exclusion did not alter the results, their data were included in the analyses. After an overnight fast, subjects reported to the laboratory for simultaneous sampling of blood and CSF (1 ml) via lumbar puncture after local anaesthesia (2 ml mepivacain-HCl 1%). BMI and waist-to-hip ratio were assessed, and body composition was measured by standard bioelectrical impedance analysis (BIA 2000-M; Data Input, Frankfurt, Germany). Insulin was determined using a commercial competitive double-antibody RIA (Pharmacia Insulin RIA 100; Pharmacia Diagnostics, Upsalla, Sweden). To increase the sensitivity of the insulin assay for the expected lower concentrations in CSF, the standard procedure was slightly modified. In brief, insulin was determined using 100 μl of CSF, 50 μl of [I]insulin diluted with buffer at a ratio of 1:3, and 50 μl of insulin antiserum diluted at a ratio of 1:2. The incubation time was increased from 2 h in the standard method to 3 h at room temperature. With this procedure, threshold sensitivity was reliably lowered to 1.8 pmol/l. Intra-assay variation was <4.5% for determination in CSF and plasma. Insulin plasma and CSF concentrations were measured in duplicate and quadruplicate, respectively. The lowest standard used was 1.8 pmol/l, which could be reliably measured with <4.5% variation. Therefore, all measures below this standard were set to 1.8 pmol/l for analysis. Thus, the actual CSF:plasma insulin ratio may even be slightly lower than reported, although analyses using the measurements below the lowest standard did not alter the results. Additionally, plasma and CSF glucose concentrations were measured (Beckman glucose analyser II; Beckman Instruments, Fullerton, CA; coefficient of variation <1.1%). Insulin resistance was estimated according to the homeostasis model assessment (HOMA) formula, with high scores denoting a high degree of insulin resistance. Bivariate correlation coefficients were determined and a stepwise multiple regression analysis was performed to detect associations Diabetologia (2006) 49:2790–2792 DOI 10.1007/s00125-006-0409-y

Cerebrospinal fluid levels of insulin in patients with Alzheimer's disease.

Some previous reports suggested a potential role of insulin in memory and in the pathophysiology of Alzheimer's disease (AD). We assessed the cerebrospinal fluid (CSF) levels of insulin in patients with AD and in age and sex-matched controls trying to elucidate whether this value could be related with the risk or severity of AD. We measured the CSF insulin levels in 27 patients with AD and 16 matched controls using a RadioImmunoanalysis method. CSF insulin levels did not differ significantly between AD-patient and control groups. These values were not correlated with age, age at onset, duration of the disease, and scores of the MiniMental State Examination in the AD group. These results suggest that CSF insulin concentrations are not related with the risk or severity of AD.

Saturable transport of insulin from plasma into the central nervous system of dogs in vivo. A mechanism for regulated insulin delivery to the brain.

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.

Effect of diet-induced obesity and experimental hyperinsulinemia on insulin uptake into CSF of the rat

We examined the hypothesis that the uptake of plasma insulin into cerebrospinal fluid (CSF) is saturable in two rat models. Dietary obese and control female Osborne Mendel rats received 24-h infusions of vehicle or insulin. CSF insulin levels in cafeteria- and chow-fed rats were comparable at all levels of plasma insulin (4.5 ± 2.8, 7.6 ± 2.4, and 23.9 ± 6.4 μU/ml in cafeteria diet vs. 4.5 ± 0.9, 6.8 ± 1.1, and 17.0 ± 4.0 μU/ ml in chow rats). CSF insulin uptake as a percentage of plasma insulin decreased with increased plasma insulin in both groups. A similar relationship was observed in Wistar rats receiving 6-day infusions of vehicle or insulin (plasma insulin = 55 ± 12 vs. 365 ± 98 μU/ml; CSF/plasma insulin ratio = 0 ± .007 vs. 0.013 ± .006, respectively). Hyperinsulinemic Wistar rats did not demonstrate decreased brain capillary insulin binding vs. vehicle-infused controls. The results suggest that a saturable transport process contributes insulin transport into CSF in normal rats and that this process is not altered by moderate diet-induced obesity or hyperinsulinemia per se.

Developmental regulation of insulin in the mammalian central nervous system

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 (∼80–90 ng/g) and early neonatal brain (∼195 ng/g) in comparison to the adult (∼32 ng/g; P < 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 circalating 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.

Seasonal changes in CSF insulin levels in marmots: insulin may not be a satiety signal for fasting in winter

Plasma insulin (PI) reportedly crosses the blood-brain barrier in mammals and acts with the central nervous system (CNS) to reduce food intake. Animals that hibernate (hibernators) eat little or no food from early winter (November) to spring (April). This lack of food intake may be due to elevated PI concentrations acting within the CNS. In this study, we determined whether hibernators have altered insulin levels within the CNS at different times during the circannual cycle of metabolism and feeding. Plasma and cerebrospinal fluid (CSF) immunoreactive insulin concentrations were measured in marmots (Marmota flaviventris) during the feeding phase of the body weight cycle and during the fasting period (hibernation). Basal plasma and CSF samples were collected in September, November, January, and April. In addition, plasma and CSF insulin levels were monitored during a 2-h intravenous infusion of glucose (20% wt/vol) that stimulated pancreatic B-cell production of insulin. During the spring feeding period, we found that as PI levels rise, so do CSF insulin concentrations. However, in fall and winter when marmots are fasting, very little insulin entered the CSF even when PI levels were significantly elevated. Furthermore, the longer the fast, the lower was the CSF insulin under both basal and infusion conditions. These results lead us to conclude that elevated CSF insulin is not a likely cause of suppressed food intake in fasting marmots.

Kinetics and specificity of insulin uptake from plasma into cerebrospinal fluid.

To characterize the relationship between insulin levels in plasma and those in cerebrospinal fluid (CSF), we studied the kinetics of both the uptake of insulin into CSF from plasma and the turnover of insulin within the CSF compartment. Sustained physiological levels of euglycemic hyperinsulinemia (plasma insulin approximately 500 pM) did not alter CSF insulin levels within the 1st h, but by 90 min a significant increase was observed (P less than 0.01). During graded hyperinsulinemic clamps (mean plasma insulin approximately 500-15,000 pM), CSF insulin rose in a dose-dependent fashion. This rise was characterized by an initial delay followed by a continuous increase for the next 150 min. We also found that after brief, high-dose intravenous insulin infusions, the t1/2 of CSF insulin was 143 +/- 7 min (means +/- SE; n = 4), similar to that of CSF turnover by bulk flow. To test the specificity of CSF insulin uptake from plasma, we compared this uptake during intravenous insulin infusions with that of proinsulin, a peptide with reduced affinity for the insulin receptor. We observed a significantly lower increment of CSF proinsulin levels over 180 min (13.6 +/- 1.6 pM; means +/- SE; n = 4) compared with that of insulin (22.4 +/- 0.6 pM; n = 4; P less than 0.01), despite plasma proinsulin levels higher than insulin (1,890 +/- 287 vs. 1,283 +/- 192 pM; P less than 0.001). When corrected for the difference in plasma levels, the uptake of insulin was fivefold greater than that of proinsulin.(ABSTRACT TRUNCATED AT 250 WORDS)