Thomas Harder

Perinatal elevation of hypothalamic insulin, acquired malformation of hypothalamic galaninergic neurons, and syndrome X-like alterations in adulthood of neonatally overfed rats

Overnutrition during critical developmental periods is suggested to be a risk factor for obesity and associated metabolic disorders in later life. Underlying mechanisms are unknown. Neuropeptides are essentially involved in the central nervous regulation of body weight. For instance, hypothalamic galanin (GAL) is a stimulator of food intake and body weight gain. To investigate long-term consequences of early postnatal overfeeding, the normal litter size of Wistar rats (n=10; controls) was reduced from day 3 to day 21 of life to only 3 pups per mother (small litters, SL; overnutrition). Throughout life, SL rats displayed hyperphagia (p<0.01), overweight (p<0.0001), hyperinsulinemia (p<0.01), impaired glucose tolerance (p<0.001), elevated triglycerides (p<0.001), and an increased systolic blood pressure (p<0.05). In adulthood, an increase of GAL-neurons in the arcuate hypothalamic nucleus (ARC) was found (p<0.001), positively correlated to body weight (p<0.001). A second experiment revealed hyperinsulinemia (p<0.001) and increased hypothalamic insulin levels (p<0.05) in SL rats during early postnatal life. Already on day 21 of life, i.e., at the end of the critical hypothalamic differentiation period, in SL rats the number of GAL-neurons was increased in the ARC (p<0.001), showing a positive correlation to body weight and insulin (p<0.05). In conclusion, neonatally acquired persisting malformation of hypothalamic galaninergic neurons, induced by early overfeeding and hyperinsulinism, might promote the development of overweight and syndrome X-like alterations during life.

Glucose tolerance and insulin secretion in children of mothers with pregestational IDDM or gestational diabetes

Summary The offspring of mothers with diabetes mellitus during pregnancy are presumed to develop altered glucose homeostasis. We analysed metabolic parameters at birth and glucose tolerance and insulin secretion during oral glucose tolerance tests at 1–9 years of age in 129 children born to mothers with pregestational insulin-dependent diabetes (IDDM) and 69 infants of gestational diabetic mothers. Newborns of IDDM mothers displayed higher insulin (p < 0.001), glucose (p < 0.05), and insulin/glucose ratios (p < 0.002) than newborns of gestational diabetic mothers. During childhood, frequencies of impaired glucose tolerance (IGT) rose in infants of IDDM mothers from 9.4 % at 1–4 years to 17.4 % at 5–9 years of age, while in children of gestational diabetic mothers an increase from 11.1 % up to 20.0 % was observed. Offspring of gestational diabetic mothers displayed higher stimulated blood glucose (p < 0.025) than infants of IDDM mothers, while children of IDDM mothers showed higher stimulated insulin (p < 0.025), accompanied by increased fasting and stimulated insulin/glucose ratios (p < 0.05 and p < 0.02, respectively). Stimulated insulin in childhood was positively correlated to insulin at birth (p < 0.05). Furthermore, insulin/glucose ratio in childhood showed a positive correlation to insulin (p < 0.01) and insulin/glucose ratio at birth (p < 0.005). In conclusion, a pathogenetic role of fetal and neonatal hyperinsulinism for the development of IGT in both groups of infants of diabetic mothers is suggested, in particular for early induction of insulin resistance in the offspring of mothers with pregestational IDDM. [Diabetologia (1997) 40: 1097–1100]

Malformations of Hypothalamic Nuclei in Hyperinsulinemic Offspring of Rats with Gestational Diabetes

Insulin is a potent modulator of central nervous development and is suggested to influence the differentiation and maturation of hypothalamic structures involved in the regulation of body weight and metabolism. Hyperinsulinemic offspring of mothers with impaired glucose tolerance during pregnancy (gestational diabetes, GD) have an increased risk to develop overweight and diabetes mellitus during life, while the underlying pathophysiological mechanisms are still unknown. To investigate the effects of perinatal hyperinsulinism on the organization of hypothalamic regulators of body weight and metabolism, GD was induced in rats by application of streptozotocin on the day of conception (25 mg/kg, i.p.). On the 21st day of life, offspring of GD rats were overweight (p < 0.05) and hyperinsulinemic (p < 0.01). Using computer-assisted morphometric measurements, significantly decreased mean areas of neuronal nuclei and neuronal cytoplasm within the paraventricular hypothalamic nucleus (PVN; p < 0.01) and the ventromedial hypothalamic nucleus (VMN; p < 0.05) were observed in GD offspring. Analysis of topographically distinct parts revealed that these alterations particularly occurred in the parvocellular part of the PVN, as well as in the anterior, central, and dorsomedial part of the VMN. No morphometric alterations were found within the lateral hypothalamic area and the dorsomedial hypothalamic nucleus. In the arcuate hypothalamic nucleus, the mean area of neuronal cytoplasm was decreased (p < 0.05), while the number of neurons expressing tyrosine hydroxylase was clearly elevated (p < 0.002). For astrocytes, a tendency towards an increased glia/neuron ratio was observed in the periventricular hypothalamic area. These observations suggest disturbed differentiation and organization of distinct hypothalamic nuclei and subnuclei, respectively, in hyperinsulinemic offspring of GD rats, possibly leading to dysfunctions of hypothalamic regulators of body weight and metabolism which might contribute to the lifelong increased risk to develop overweight and diabetogenic disturbances.

Hypothalamic neuropeptide Y levels in weaning offspring of low-protein malnourished mother rats.

Maternal low-protein malnutrition during gestation and lactation (LP) is an animal model frequently used for the investigation of long-term deleterious consequences of perinatal growth retardation. Hypothalamic neuropeptides are decisively involved in the central nervous regulation of body weight and metabolism. We investigated neuropeptide Y (NPY) in distinct hypothalamic nuclei in the offspring of LP mother rats at the end of the critical hypothalamic differentiation period (20th day of life). Weanling LP offspring were underweight (P< 0.001) and hypoinsulinaemic (P< 0.05), while leptin levels were unchanged. NPY was significantly increased in the paraventricular hypothalamic nucleus (PVN) (P< 0.01) and lateral hypothalamic area (P< 0.05) in LP offspring. In contrast, NPY was unchanged in the ventromedial hypothalamic nucleus (VMN). These observations indicate a leptin-independent stimulation of the orexigenic ARC-PVN axis in undernourished LP rats at weaning. Furthermore a disturbed NPYergic regulation of the VMN is suggested, possibly contributing to alterations of the hypothalamic regulation of body weight and metabolism in LP offspring during life.

Syndrome X-like alterations in adult female rats due to neonatal insulin treatment.

Hypothalamic structures are decisively involved in the regulation of body weight and metabolism. In syndrome X, complex metabolic alterations are present, which in women are found to be associated with disturbances of reproductive function and altered androgen levels. In previous experiments in rats, it was shown that a temporary intrahypothalamic hyperinsulinism during early life predisposes to overweight and diabetogenic disturbances later in life, associated with disorganization of hypothalamic regulatory centers. To investigate the possible long-term consequences of elevated peripheral insulin levels during ontogenesis, the following experiment was performed. Newborn female Wistar rats were treated during neonatal life with daily subcutaneous injections of long-acting insulin ([IRI group] 0.3 IU on days 8 and 9 of life and 0.1 IU on days 10 and 11 of life), whereas control animals (CO) received daily NaCl injections. This temporary exposure to increased insulin levels during a critical developmental period resulted in an increased body weight gain including juvenile life and adulthood (P < .01), accompanied by hyperinsulinemia (P < .01), impaired glucose tolerance (P < .05), and increased systolic blood pressure in adulthood (P < .025). No significant alterations were detected either in cyclicity and fertility or in the levels of testosterone, androstenedione, or dehydroepiandrosterone (DHEA) in IRI rats. Morphometric evaluation of hypothalamic nuclei showed a reduced numerical density of neurons (P < .025) and a decreased neuronal volume density (P < .025) within the ventromedial hypothalamic nucleus (VMN) of the IRI rats, whereas the antagonistic lateral hypothalamic area (LHA) was morphometrically unchanged. Newborn offspring of IRI rats (F1 generation) were overweight (P < .05) and had an increased pancreatic insulin concentration (P < .02). In conclusion, perinatal hyperinsulinism seems to predispose to the later development of syndrome X-like changes in female rats, possibly due to impaired organization of hypothalamic regulators of body weight and metabolism.

Increased number of galanin-neurons in the paraventricular hypothalamic nucleus of neonatally overfed weanling rats

Perinatal overfeeding is a risk factor for overweight and diabetes during life. Underlying pathophysiological mechanisms are unclear. The peptide galanin is suggested to stimulate food intake by acting within the paraventricular hypothalamic nucleus (PVN). In early postnatally overfed rats overweight and hyperinsulinemia were observed, accompanied by an increased number of galanin-positive neurons in the PVN at weaning. Our results might indicate malformation of hypothalamic galaninergic neurons due to neonatal overfeeding and hyperinsulinism, respectively, in rats.

Reduction of cholecystokinin-8S-neurons in the paraventricular hypothalamic nucleus of neonatally overfed weanling rats

Cholecystokinin (CCK) is suggested to be involved, e.g. in the central nervous modulation of food intake, possibly by acting within specific hypothalamic nuclei. Perinatal overnutrition predisposes to permanent obesity and hyperphagia, while underlying mechanisms are unclear. By reducing the litter size from the 3rd to 21st day of life, early overnutrition was induced in newborn rats. At weaning, clear overweight (P < 0.001), hyperglycaemia (P < 0.05), hyperinsulinaemia (P < 0.001), and insulin resistance (P < 0.001) occured. These early signs of obesity were associated with a significantly decreased number of CCK-positive neurons in the paraventricular hypothalamic nucleus (P < 0.002). In conclusion, due to neonatal overfeeding malformation of CCKergic neurons at the end of the critical hypothalamic differentiation period occurs. Long-term consequences on CCK-related neuroendocrine regulations could be suggested, including those affecting food intake and body weight gain.

Alterations of hypothalamic catecholamines in the newborn offspring of gestational diabetic mother rats

Catecholamines are essential organizers of the developing brain. Throughout life, they are involved, e.g., in the regulation of body weight and metabolism by specific hypothalamic nuclei, which are suggested to be highly vulnerable to maternal gestational hyperglycemia. By application of streptozotocin (30 mg/kg, i.p.) gestational diabetes (GD) was induced in female rats. On the 1st day of life, male GD offspring were underweight (P<0.05) and hyperglycemic (P<0.05), while on the 21st day of life decreased body weight (P<0.001) and elevated pancreatic insulin (P<0.01) were observed. Using HPLC with electrochemical detection, hypothalamic catecholamines were determined in the newborns, and quantitative immunocytochemistry for tyrosine hydroxylase (TH) was performed. At birth, a tendency towards increased levels of norepinephrine (NE) and dopamine (DA) in the whole hypothalami of GD offspring was observed. In the 21-day-old offspring of GD mothers, NE was significantly increased in the ventromedial hypothalamic nucleus (VMN; P<0.05) and the lateral hypothalamic area (LHA; P<0.05), while DA was significantly elevated in the paraventricular hypothalamic nucleus (PVN; P<0.05) and the LHA (P<0.05). The NE/DA-ratio was found to be decreased in the PVN of GD offspring (P<0.01). Moreover, numerical density of TH-positive neurons was clearly increased within the parvocellular division of the PVN (P<0.0001) as well as in the periventricular hypothalamic area (PER; P<0.05). These data suggest specific alterations of catecholaminergic systems within hypothalamic regulators of body weight and metabolism during early development in the offspring of gestational diabetic mother rats.

Maternal and Paternal Family History of Diabetes in Women with Gestational Diabetes or Insulin-Dependent Diabetes mellitus Type I

Animal studies have shown that prenatal exposure to a diabetic intrauterine milieu leads to an increased risk in the female offspring of developing gestational diabetes (GD). In the present study, the family history of non-insulin-dependent diabetes mellitus type II (NIDDM) and insulin-dependent diabetes mellitus type I (IDDM) was evaluated in 106 women with GD, as compared to 189 women with IDDM. In GD patients, the prevalence of diabetes was significantly greater in mothers than in fathers (p = 0.03). This was mainly due to a greater prevalence of NIDDM in the mothers (p = 0.05). Furthermore, a significant aggregation of NIDDM was also observed in the maternal-grandmaternal line of GD women, as compared to the paternal-grandpaternal side (p = 0.02). In patients with IDDM no significant difference concerning the prevalence of any type of diabetes between mothers and fathers was observed. In conclusion, an aggregation of NIDDM in mothers and grandmothers of women with GD is reported here. A history of NIDDM on the maternal side of pregnant women should be considered as a particular risk factor for GD and, hence, for intergenerative transmission of NIDDM, which therefore might be prevented, at least in part, by strict avoidance of GD.

Pancreatic islet transplantation in diabetic pregnant rats prevents acquired malformation of the ventromedial hypothalamic nucleus in their offspring

Exposure to a diabetic intrauterine environment leads to diabetogenic disturbances throughout later life in rats. This is accompanied by a fetally acquired dysplasia of the ventromedial hypothalamic nucleus (VMN) which is decisively involved in the regulation of metabolism. We investigated whether malformation of the VMN is preventable by normalization of gestational hyperglycaemia. Correction of hyperglycaemia in pregnant streptozotocin-diabetic rats was achieved by pancreatic islet transplantation. The number of neurons in the VMN was significantly reduced in adult offspring of non-treated, sham-transplanted mother rats (P<0.05), but did not differ between offspring of islet-transplanted mother rats and offspring of control mothers. In conclusion, prevention of VMN malformation in offspring of islet-transplanted diabetic mothers might be co-responsible for normalization of their glucose homeostasis during life.