Anneke Alkemade

Hypothalamic thyroid hormone feedback in health and disease

The role of the human hypothalamus in the neuroendocrine response to illness has only recently begun to be explored. Extensive changes in the hypothalamus-pituitary-thyroid (HPT) axis occur within the framework of critical illness. The best-documented change in the HPT axis is a decrease in serum concentrations of the biologically active thyroid hormone triiodothyronine (T3). From studies in post-mortem human hypothalamus it appeared that low serum T3 and thyrotropin (TSH) during illness (nonthyroidal illness, NTI) are paralleled by decreased thyrotropin-releasing hormone (TRH)mRNA expression in the hypothalamic paraventricular nucleus (PVN), pointing to a major alteration in HPT axis setpoint regulation. A strong decrease in TRHmRNA expression is also present in the PVN of patients with major depression as well as in glucocorticoid-treated patients. By inference, hypercortisolism in hospitalized patients with severe depression or in critical illness may induce down-regulation of the HPT axis at the level of the hypothalamus. In order to start defining the determinants and mechanisms of these setpoint changes in various clinical conditions, it is important to note that an increasing number of hypothalamic proteins appears to be involved in central thyroid hormone metabolism. In recent studies, we have investigated the distribution and expression of thyroid hormone receptor (TR) isoforms, type 2 and type 3 deiodinase (D2 and D3), and the thyroid hormone transporter monocarboxylate transporter 8 (MCT8) in the human hypothalamus by a combination of immunocytochemistry, mRNA in situ hybridization and enzyme activity assays. Both D2 and D3 enzyme activities are detectable in the mediobasal hypothalamus. D2 immunoreactivity is prominent in glial cells of the infundibular nucleus/median eminence region and in tanycytes lining the third ventricle. Combined D2, D3, MCT8 or TR immunocytochemistry and TRHmRNA in situ hybridization indicates that D3, MCT8 and TRs are all expressed by TRH neurons in the PVN, whereas D2 is not. Taken together, these results suggest that the prohormone thyroxine (T4) is taken up in glial cells that convert T4 into the biologically active T3 via the enzyme D2; T3 is subsequently transported to TRH producing neurons in the PVN where it may bind to TRs and/or may be degraded into inactive iodothyronines by D3. This model for thyroid hormone action in the human hypothalamus awaits confirmation in future experimental studies.

Hypothalamic Neuropeptide Y (NPY) Controls Hepatic VLDL-Triglyceride Secretion in Rats via the Sympathetic Nervous System

Excessive secretion of triglyceride-rich very low-density lipoproteins (VLDL-TG) contributes to diabetic dyslipidemia. Earlier studies have indicated a possible role for the hypothalamus and autonomic nervous system in the regulation of VLDL-TG. In the current study, we investigated whether the autonomic nervous system and hypothalamic neuropeptide Y (NPY) release during fasting regulates hepatic VLDL-TG secretion. We report that, in fasted rats, an intact hypothalamic arcuate nucleus and hepatic sympathetic innervation are necessary to maintain VLDL-TG secretion. Furthermore, the hepatic sympathetic innervation is necessary to mediate the stimulatory effect of intracerebroventricular administration of NPY on VLDL-TG secretion. Since the intracerebroventricular administration of NPY increases VLDL-TG secretion by the liver without affecting lipolysis, its effect on lipid metabolism appears to be selective to the liver. Together, our findings indicate that the increased release of NPY during fasting stimulates the sympathetic nervous system to maintain VLDL-TG secretion at a postprandial level.

Functional neuroanatomy of thyroid hormone feedback in the human hypothalamus and pituitary gland

A major change in thyroid setpoint regulation occurs in various clinical conditions such as critical illness and psychiatric disorders. As a first step towards identifying determinants of these setpoint changes, we have studied the distribution and expression of thyroid hormone receptor (TR) isoforms, type 2 and type 3 deiodinase (D2 and D3), and the thyroid hormone transporter monocarboxylate transporter 8 (MCT8) in the human hypothalamus and anterior pituitary. Although the post-mortem specimens used for these studies originated from patients who had died from many different pathologies, the anatomical distribution of these proteins was similar in all patients. D2 enzyme activity was detectable in the infundibular nucleus/median eminence (IFN/ME) region coinciding with local D2 immunoreactivity in glial cells. Additional D2 immunostaining was present in tanycytes lining the third ventricle. Thyrotropin-releasing hormone (TRH) containing neurons in the paraventricular nucleus (PVN) expressed MCT8, TRs as well as D3. These findings suggest that the prohormone thyroxine (T4) is taken up in hypothalamic glial cells that convert T4 into the biologically active triiodothyronine (T3) via the enzyme D2, and that T3 is subsequently transported to TRH producing neurons in the PVN. In these neurons, T3 may either bind to TRs or be metabolized into inactive iodothyronines by D3. By inference, local changes in thyroid hormone metabolism resulting from altered hypothalamic deiodinase or MCT8 expression may underlie the decrease in TRH mRNA reported earlier in the PVN of patients with critical illness and depression. In the anterior pituitary, D2 and MCT8 immunoreactivity occurred exclusively in folliculostellate (FS) cells. Both TR and D3 immunoreactivity was observed in gonadotropes and to a lesser extent in thyrotropes and other hormone producing cell types. Based upon these neuroanatomical findings, we propose a novel model for central thyroid hormone feedback in humans, with a pivotal role for hypothalamic glial cells and pituitary FS cells in processing and activation of T4. Production and action of T3 appear to occur in separate cell types of the human hypothalamus and anterior pituitary.

Thyroid hormone is required for hypothalamic neurons regulating cardiovascular functions

Thyroid hormone is well known for its profound direct effects on cardiovascular function and metabolism. Recent evidence, however, suggests that the hormone also regulates these systems indirectly through the central nervous system. While some of the molecular mechanisms underlying the hormones central control of metabolism have been identified, its actions in the central cardiovascular control have remained enigmatic. Here, we describe a previously unknown population of parvalbuminergic neurons in the anterior hypothalamus that requires thyroid hormone receptor signaling for proper development. Specific stereotaxic ablation of these cells in the mouse resulted in hypertension and temperature-dependent tachycardia, indicating a role in the central autonomic control of blood pressure and heart rate. Moreover, the neurons exhibited intrinsic temperature sensitivity in patch-clamping experiments, providing a new connection between cardiovascular function and core temperature. Thus, the data identify what we believe to be a novel hypothalamic cell population potentially important for understanding hypertension and indicate developmental hypothyroidism as an epigenetic risk factor for cardiovascular disorders. Furthermore, the findings may be beneficial for treatment of the recently identified patients that have a mutation in thyroid hormone receptor α1.

Glucocorticoid Signaling in the Arcuate Nucleus Modulates Hepatic Insulin Sensitivity

Glucocorticoid receptors are highly expressed in the hypothalamic paraventricular nucleus (PVN) and arcuate nucleus (ARC). As glucocorticoids have pronounced effects on neuropeptide Y (NPY) expression and as NPY neurons projecting from the ARC to the PVN are pivotal for balancing feeding behavior and glucose metabolism, we investigated the effect of glucocorticoid signaling in these areas on endogenous glucose production (EGP) and insulin sensitivity by local retrodialysis of the glucocorticoid receptor agonist dexamethasone into the ARC or the PVN, in combination with isotope dilution and hyperinsulinemic–euglycemic clamp techniques. Retrodialysis of dexamethasone for 90 min into the ARC or the PVN did not have significant effects on basal plasma glucose concentration. During the hyperinsulinemic–euglycemic clamp, retrodialysis of dexamethasone into the ARC largely prevented the suppressive effect of hyperinsulinemia on EGP. Antagonizing the NPY1 receptors by intracerebroventricular infusion of its antagonist largely blocked the hepatic insulin resistance induced by dexamethasone in the ARC. The dexamethasone-ARC–induced inhibition of hepatic insulin sensitivity was also prevented by hepatic sympathetic denervation. These data suggest that glucocorticoid signaling specifically in the ARC neurons modulates hepatic insulin responsiveness via NPY and the sympathetic system, which may add to our understanding of the metabolic impact of clinical conditions associated with hypercortisolism.

Expression of thyroid hormone transporters in the human hypothalamus.

CONTEXT Transport of thyroid hormone across the plasma membrane is required for proper thyroid hormone action and metabolism. Several specific thyroid hormone transporters have been identified capable of facilitating uptake and/or efflux of thyroid hormones. Monocarboxylate transporter (MCT)-8, MCT10, and organic anion transporting polypeptide 1C1 (OATP1C1) are the best-characterized specific thyroid hormone transporters to date. OBJECTIVE Our earlier studies in the human hypothalamus have shown that MCT8 is present in neurons of the hypothalamic paraventricular nucleus (PVN) and infundibular nucleus (IFN) and in tanycytes. We hypothesized that also MCT10 and OATP1C1 are present in specific areas of the human hypothalamus. DESIGN We studied postmortem brain samples of patients with known serum thyroid hormone levels using immunocytochemistry to investigate the distribution of MCT10 and OATP1C1 in the hypothalamus. RESULTS We found strong neuronal MCT10 immunocytochemical staining in a number of hypothalamic nuclei, including the PVN, IFN, and supraoptic nucleus. Intense staining was also observed in neurons of the lateral hypothalamus including the perifornical area. OATP1C1 immunoreactivity was present in glial cells throughout the hypothalamus. In addition, staining was present in capillary walls and in neurons of the PVN, IFN, and supraoptic nucleus. CONCLUSION The strong expression of MCT10 and OATP1C1 in the human hypothalamus indicates a possible role in the regulation of the hypothalamus-pituitary-thyroid axis.

Melanocortin 4 receptor distribution in the human hypothalamus

OBJECTIVE The melanocortin 4 receptor (MC4R) is an essential regulator of energy homeostasis and metabolism, and MC4R mutations represent the most prevalent monogenetic cause of obesity in humans known to date. Hypothalamic MC4Rs in rodents are well characterized in neuroanatomical and functional terms, but their expression pattern in the human hypothalamus is unknown. DESIGN AND METHODS To determine the topographic distribution and identity of cells expressing MC4R mRNA in the human hypothalamus, locked nucleic acid in situ hybridization was performed on nine human postmortem hypothalami. In addition, co-expression of MC4R with glial fibrillary acidic protein (GFAP), vasopressin/oxytocin (AVP/OXT), corticotropin-releasing hormone (CRH), neuropeptide Y (NPY), agouti-related protein (AgRP), and α-melanocyte stimulating hormone (α-MSH) was examined. RESULTS Most intense MC4R mRNA expression was present in the paraventricular nucleus (PVN), the supraoptic nucleus (SON), and the nucleus basalis of Meynert. Most MC4R-positive cells in the SON also expressed AVP/OXT. Co-expression with AVP/OXT in the PVN was less abundant. We did not observe co-expression of MC4R mRNA and GFAP, CRH, NPY, AgRP, or α-MSH. However, fiber-like staining of NPY, AgRP, and α-MSH was found adjacent to MC4R-positive cells in the PVN. CONCLUSION Expression of MC4R mRNA in the human hypothalamus is widespread and in close approximation to endogenous MC4R binding partners AgRP and α-MSH.

Multi-modal ultra-high resolution structural 7-Tesla MRI data repository

Structural brain data is key for the understanding of brain function and networks, i.e., connectomics. Here we present data sets available from the ‘atlasing of the basal ganglia (ATAG)’ project, which provides ultra-high resolution 7 Tesla (T) magnetic resonance imaging (MRI) scans from young, middle-aged, and elderly participants. The ATAG data set includes whole-brain and reduced field-of-view MP2RAGE and T2*-weighted scans of the subcortex and brainstem with ultra-high resolution at a sub-millimeter scale. The data can be used to develop new algorithms that help building high-resolution atlases both relevant for the basic and clinical neurosciences. Importantly, the present data repository may also be used to inform the exact positioning of electrodes used for deep-brain-stimulation in patients with Parkinson’s disease and neuropsychiatric diseases.

AgRP and NPY Expression in the Human Hypothalamic Infundibular Nucleus Correlate with Body Mass Index, Whereas Changes in αMSH Are Related to Type 2 Diabetes

CONTEXT Rodent data show that altered hypothalamic signaling contributes to the development of obesity and insulin resistance. OBJECTIVE To determine differences in hypothalamic expression levels of neuropeptide Y (NPY), agouti-related peptide (AgRP), and αMSH in the infundibular nucleus, the human equivalent of the arcuate nucleus, in relation to body mass index (BMI). In addition, the expression in the infundibular nucleus of eight subjects diagnosed with type 2 diabetes was measured to determine possible interference of type 2 diabetes with the association observed between neuropeptides and BMI. DESIGN We studied AgRP, NPY, and αMSH expression by means of quantitative immunocytochemistry in postmortem hypothalami of 30 subjects with known BMI. In separate experiments, we compared neuropeptide expression in eight subjects with type 2 diabetes with eight matched controls. RESULTS We found that AgRP immunoreactivity showed a U-shaped correlation with BMI. No evidence was found for possible influences of corticosteroid treatment. NPY immunoreactivity was significantly lower in overweight and obese subjects. αMSH did not correlate with BMI but was significantly lower in subjects with type 2 diabetes compared with controls. By contrast, NPY and AgRP expression was not affected in type 2 diabetes. CONCLUSION Our results indicate that the expression of AgRP and NPY are correlated with body weight changes, rather than the presence of type 2 diabetes, whereas changes in αMSH immunoreactivity are related to the presence of type 2 diabetes, indicating separate hypothalamic mechanisms.

High calorie diet triggers hypothalamic angiopathy

Obesity, type 2 diabetes, and related diseases represent major health threats to modern society. Related pathophysiology of impaired neuronal function in hypothalamic control centers regulating metabolism and body weight has been dissected extensively and recent studies have started focusing on potential roles of astrocytes and microglia. The hypothalamic vascular system, however, which maintains the microenvironment necessary for appropriate neuronal function, has been largely understudied. We recently discovered that high fat/high sucrose diet exposure leads to increased hypothalamic presence of immunoglobulin G (IgG1). Investigating this phenomenon further, we have discovered a significant increase in blood vessel length and density in the arcuate nucleus (ARC) of the hypothalamus in mice fed a high fat/high sucrose diet, compared to matched controls fed standard chow diet. We also found a clearly increased presence of α-smooth muscle actin immunoreactive vessels, which are rarely present in the ARC and indicate an increase in the formation of new arterial vessels. Along the blood brain barrier, an increase of degenerated endothelial cells are observed. Moreover, such hypothalamic angiogenesis was not limited to rodent models. We also found an increase in the number of arterioles of the infundibular nucleus (the human equivalent of the mouse ARC) in patients with type 2 diabetes, suggesting angiogenesis occurs in the human hypothalamus of diabetics. Our discovery reveals novel hypothalamic pathophysiology, which is reminiscent of diabetic retinopathy and suggests a potential functional involvement of the hypothalamic vasculature in the later stage pathogenesis of metabolic syndrome.