Ligation of the uterine artery and early postnatal food restriction - animal models for growth retardation.

Intrauterine growth restriction (IUGR) is one of the major causes of short stature in child- and adulthood. The cause of IUGR is unknown, however, an impaired uteroplacental function during the second half of human pregnancy might be an important factor, by affecting the programming of somatotropic axis and leading to postnatal growth failure into adulthood. Two rat models with perinatally induced growth retardation were used to examine the long-term effects of perinatal insults on growth. IUGR rats were prepared from pregnant dams, with a bilateral uterine artery ligation at day 17 of their pregnancy. Since the rat is relatively immature at birth, an early postnatal food restriction model was included as another model to broaden the time window of sensitive period of organogenesis. An individual growth curve was calculated of each animal (n = 813). From these individual growth curves the predicted growth curve for each experimental group was calculated by multilevel analysis. The proposed mathematical model allows us to estimate the growth potentials of these rat models with precision and could provide basic information to investigate the relationships among a number of other variables in future studies. Furthermore, we concluded that both pre- and early postnatal malnutrition leads to irreversible slowing down of postnatal growth.

Decreased galanin mRNA levels in growth hormone-releasing hormone neurons after perinatally induced growth retardation.

Intrauterine growth retardation (IUGR) is associated with persistent postnatal growth retardation accompanied by dysfunction of the hypothalamic components of the growth hormone (GH) axis. At the adult stage, this is reflected by increased somatostatin (SS) and decreased neuropeptide Y (NPY) mRNA levels, whereas the GH-releasing hormone (GHRH) mRNA levels are normal and the output of GH remains unchanged. To extend our insight into the hypothalamic control of GH secretion in growth retarded rats, we determined galanin (GAL) mRNA levels at the adult stage of perinatally malnourished (i.e. IUGR and early postnatally food restricted) rats. Analyses included comparison of GAL mRNA levels in GHRH neurons in perinatally malnourished adult rats using a semi-quantitative double labeling in situ hybridization technique. We report that IUGR is accompanied by a 60% decrease in GAL mRNA levels in all GHRH neurons in the male IUGR group whereas a tendency towards a decrease was observed in the male early postnatally food restricted (FR) group. These effects became more pronounced when the analysis was restricted to GHRH neurons coexpressing GAL mRNA i.e. decreased GAL mRNA levels were seen in both male and female IUGR rats and in FR males. These data show that GAL mRNA levels in GHRH neurons are persistently decreased after perinatal malnutrition. Taking these results together with our previous data on SS, NPY and GHRH mRNA levels, we can conclude that IUGR leads to a reprogramming of the hypothalamic regulation of GH secretion.

Persistent changes in somatostatin and neuropeptide Y mRNA levels but not in growth hormone-releasing hormone mRNA levels in adult rats after intrauterine growth retardation.

A reduction in the availability of oxygen and nutrients across the placenta in the last trimester of pregnancy may lead to intrauterine growth retardation (IUGR) which, in turn, may cause a persistent postnatal growth failure. However, it is unknown whether this persistent growth retardation is centrally mediated through alterations in the components of the growth hormone (GH)-axis. We tested the hypothesis that alterations in the development of the central components of the GH-axis contribute to the persistent growth failure observed after experimentally induced IUGR or early postnatal food restriction (FR) in the rat. Using semi-quantitative in situ hybridization, we compared somatostatin (SS), GH-releasing hormone (GHRH) and neuropeptide Y (NPY) mRNA levels in adult rats experimentally subjected to IUGR or FR. We report that IUGR increased the expression of SS mRNA in the periventricular nucleus (PeN) of adult male and female rats by 128% and 153% respectively, did not alter the expression of GHRH mRNA in the arcuate nucleus (ARC) and decreased the NPY mRNA expression in the ARC by 73% in males and 61% in females, whereas in the FR group no changes in the expression of these mRNAs were observed. These data show that the timing of malnutrition or the presence of the placenta is important for the long-term alterations since the effects only occurred in the prenatally induced growth retardation and not in the early postnatally induced growth retardation group.

Effects of intrauterine and early postnatal growth restriction on hypothalamic somatostatin gene expression in the rat.

In the human, intrauterine growth retardation (IUGR) can result in persistent postnatal growth failure, which may be attributable, in part, to abnormal GH secretion. Whether putative alterations in GH secretion are the result of abnormalities intrinsic to the pituitary or reflect changes in the production of GH-releasing hormone or somatostatin (SS) is unknown. We tested the hypothesis that growth failure associated with IUGR or early postnatal food restriction (FR) is caused by a central defect in hypothalamic SS gene expression. Both models displayed persistent growth failure postnatally without any catch-up growth. We measured levels of SS mRNA levels in rats experimentally subjected to IUGR or FR. SS mRNA levels were measured by semiquantitative in situ hybridization throughout development. Levels of SS mRNA in the periventricular nucleus were significantly higher in both male and female IUGR rats in the juvenile and adult stages compared with matched controls (p < or = 0.05). FR was associated with higher SS mRNA levels only in neonatal female rats (p < or = 0.05). These results suggest that intrauterine malnutrition induces a persistent increase in the expression of SS mRNA in the periventricular nucleus, whereas early postnatal FR results in only a transient increase in SS gene expression. Because IGF-I levels were normal in juvenile IUGR and FR rats, central dysregulation of SS neurons does not appear to be the cause of early postnatal growth failure in either model. However, these observations are consistent with the hypothesis that nutritional stress at critical times during development can have persistent and potentially irreversible effects on organ function.

Transgenic expression of B-50/GAP-43 in mature olfactory neurons triggers downregulation of native B-50/GAP-43 expression in immature olfactory neurons.

The adult mammalian olfactory neuroepithelium is an unusual neural tissue, since it maintains its capacity to form new neurons throughout life. Newly formed neurons differentiate in the basal layers of the olfactory neuroepithelium and express B-50/GAP-43, a protein implicated in neurite outgrowth. During maturation these neurons migrate into the upper portion of the epithelium, upregulate expression of olfactory marker protein (OMP) and concomitantly downregulate the expression of B-50/GAP-43. Transgenic mice that exhibit OMP-promoter directed expression of B-50/GAP-43 in mature olfactory neurons display an unexpected decrease in the complement of B-50/GAP-43-positive cells in the lower region of the olfactory epithelium [A.J.G.D. Holtmaat, P.A. Dijkhuizen, A.B. Oestreicher, H. J. Romijn, N.M.T. Van der Lugt, A. Berns, F.L. Margolis, W.H. Gispen, J. Verhaagen, Directed expression of the growth-associated protein B-50/GAP-43 to olfactory neurons in transgenic mice results in changes in axon morphology and extraglomerular growth, J. Neurosci. 15 (1995) 7953-7965]. We have investigated whether the decrement in B-50/GAP-43-positive cells in this region was due to a dislocation of the immature neurons to other regions of the olfactory epithelium or to a downregulation of B-50/GAP-43 synthesis in these immature neurons. In eight of nine independent transgenic mouse lines that express the transgene in different numbers of olfactory neurons, a decline in the number of B-50/GAP-43-expressing neurons in the basal portion of the olfactory neuroepithelium was observed, both at the protein level and the mRNA level. An alternative marker for immature cells, a juvenile form of tubulin, was normally expressed in this location, indicating that the olfactory epithelium of OMP-B-50/GAP-43 transgenic mice contains a normal complement of immature olfactory neurons and that most of these neurons display a downregulation of B-50/GAP-43 expression.