Development of a Method for the Determination of Pulsatile Growth Hormone Secretion in Mice

Abstract

Measures of pulsatile GH secretion require frequent collection and analysis of blood samples at regular intervals. Due to blood volume constraints, repeat measures of circulating levels of GH in mice remain challenging. Consequently, few observations exist in which the pulsatile pattern of GH secretion in mice have been characterized. To address this, we developed a technique for the collection and analysis of circulating levels of GH at regular and frequent intervals in freely moving mice. This was achieved through the development of a sensitive assay for the detection of GH in small (2 μl) quantities of whole blood. The specificity and accuracy of this assay was validated following guidelines established for single-laboratory validation as specified by the International Union of Pure and Applied Chemistry. We incorporated an established method for tail-clip blood sample collection to determine circulating levels of GH secretion in 36 whole blood samples collected consecutively over a period of 6 h. Resulting measures were characterized by peak secretion periods and interpulse stable baseline secretion periods. Periods characterized by elevated whole blood GH levels consisted of multicomponent peaks. Deconvolution analysis of resulting measures confirmed key parameters associated with pulsatile GH secretion. We show a striking decrease in pulsatile GH secretion in mice after 12-18 h of fasting. This model is necessary to characterize the pulsatile profile of GH secretion in mice and will significantly contribute to current attempts to clarify mechanisms that contribute to the regulation of GH secretion.

Phosphorylation of the adaptor protein SH2B1βregulates its ability to enhance growth hormone (GH)-dependent macrophage motility

“…1B). These data indicate that GH is able to induce directed macrophage movement (chemotaxis) at concentrations of GH well within the physiological range for rodent plasma GH [pulsatile peak levels of 200-300 ng/ml (Edén, 1979;Steyn et al, 2011) and ghrelin-stimulated levels of GH up to 600 ng/ml (Morozumi et al, 2011)]. Based on these results, GH was added only to the lower chamber of the transwell plate for subsequent migration assays.…”

Enhanced responsiveness of Ghsr Q343X rats to ghrelin results in enhanced adiposity without increased appetite

“…After centrifugation, plasma was acidified with HCl (0.1 M final) and stored at −80°C. Plasma GH concentrations were determined by a double antibody EIA assay using materials supplied by the National Hormone and Peptide Program (NHPP) as 20 previously described (49). Insulin concentrations were determined by ELISA (Mercodia) and rat acyl ghrelin and des-acyl ghrelin concentrations were measured with ELISA kits (Bertin Pharma).…”

The effect of hexarelin in streptozotocin (STZ)-induced rat model through protective actions on islet beta cells and cardiomyocytes

“…The method has been previously described [253]. Animals had free access to food and water through blood collection.…”

“…To determine GH concentration, a sensitive sandwich ELISA had been developed and validated (as shown in Figure 2.1) [253]. In general, a 96-well plate was coated by monkey anti-rat antibody …”