Effect of gsp oncogene on somatostatin receptor subtype 1 and 2 mRNA levels in GHRH-responsive GH3 cells.

Growth hormone releasing hormone (GHRH) signals via G protein-coupled receptors (GHRH-R) to enhance intracellular Galphas/adenylyl cyclase/cAMP signaling, which in turn has positive effects on GH synthesis and release, as well as proliferation of the GH-producing cells of the anterior pituitary gland. Some GH-producing pituitary tumors express a constitutively active mutant form of Galphas (gsp oncogene). It has been reported that these tumors are more responsive to octreotide therapy. In this study we used a rat GH-producing cell line (GH3) stably transfected with the human GHRH-R cDNA (GH3-GHRHR cells) as a model to study the effects of gsp oncogene on somatostatin (SRIH) receptor subtype 1 and 2 (sst1 and sst2) mRNA levels. Transient transfection of gsp oncogene in GH3-GHRHR cells for 48 h increased intracellular cAMP levels and GH release. Phosphodiesterase (PDE) 4, sst1 and sst2 mRNA levels were increased by G protein mutation as assessed by real-time RT-PCR. Increased PDE mRNA levels in gsp-transfected cells may be a compensatory mechanism to the constitutive activation of cAMP-dependent pathway by G protein mutation and is consistent with reports of higher PDE expression in human pituitary tumor that express gsp. Our data suggest that higher expression of sst1 and sst2 mRNA induced by the gsp oncogene may be a mechanism by which gsp-positive tumors show a greater response to SRIH. GH3 cells permanently transfected with GHRH-R can be used for in vitro studies of actions of GHRH.

Acute hyperglycemia and activation of the beta-adrenergic system do not exhibit synergistic inhibitory actions on thyrotropin-releasing hormone (TRH)-induced thyroid stimulating hormone (TSH) secretion.

Thyrotropin-releasing hormone (TRH)-stimulated thyroid stimulating hormone (TSH) response in normal subjects is suppressed by oral glucose administration. Pharmacologic studies indicate that this suppressive action of glucose is mediated by an increase in hypothalamic somatostatin (SRIH) tone. Since activation of the beta-adrenergic system also suppresses basal TSH secretion by enhancing SRIH release we sought to determine whether isoproterenol alters the suppression of TRH-induced TSH response induced by the stimulation of glucose. Four tests were performed in seven healthy young men: Test 1: 200 microg TRH (iv) at 0 min; Test 2: 100 g oral glucose at -30 min and TRH at 0 min; Test 3: TRH at 0 min with isoproterenol (0.012 microg/kg, iv) infused continuously; Test 4: oral glucose at -30 min, TRH at 0 min with isoproterenol infused continuously. Pretreatment with glucose significantly suppressed TRH-induced TSH secretion. Isoproterenol infusion also suppressed the TRH-induced TSH secretion, but it did not enhance the inhibitory action of glucose on TSH secretion. The degree of suppression induced by glucose was significantly higher than that achieved by isoproterenol. These data suggest that combined administration of glucose and isoproterenol does not exhibit synergistic inhibitory actions on TRH-stimulated TSH secretion, and that the glucose-TRH test could be used for the evaluation of the hypothalamic somatostatinergic activity.

Somatostatin (SRIF) receptor subtype 2 and 5 gene expression in growth hormone-secreting pituitary adenomas: the relationship with endogenous srif activity and response to octreotide.

To investigate the potential pathophysiologic role of human SRIF receptor gene expression in GH-secreting adenomas in acromegalic patients, we studied the relationship between the SRIF receptor gene expression, endogenous SRIF activity and exogenous response to octreotide in 16 acromagalic patients. Hypothalamic somatostatinergic activity (HSA) was assessed by glucose-induced suppression of TRH-stimulated TSH secretion. As an indicator of somatotrope sensitivity to HSA, glucose-induced suppression of TRH-stimulated GH secretion was determined. For the acute octreotide response, a 100 microg bolus of octreotide was injected intravenously and GH was measured hourly for 6 hr. Pituitary tumor SRIF receptor subtype 2 and 5 (sst2 and sst5) mRNA levels were measured by real-time RT-PCR. Gsp oncogene was also detected by direct PCR sequencing. Sst2 and sst5 mRNA levels were detected in all tumors. Sst2 mRNA levels positively correlated with that of sst5. Sst2 and sst5 mRNA levels did not show any correlation with basal GH values (nadir or peak). Expression of sst2, but not sst5, showed a positive correlation with the GH response to HSA, while the octreotide response positively correlated with the sum of sst2 and sst5 mRNA levels. Individuals with gsp-positive tumors were more responsive to octreotide than those with gsp-negative tumors but sst2 and sst5 mRNA levels did not differ between these two groups. These results suggest common transcriptional and/or post-transcriptonal regulatory mechanisms for these SRIF receptor subtypes within GH-secreting pituitary adenomas. The functional observations suggest that the degree (or level) of sst2 and sst5 expression is critical for the ultimate GH response of somatotropinomas to endogenous SRIF tone and exogenous SRIF analogue therapy. However, sst2 and sst5 mRNA levels are not the only factors mediating the response to SRIF.

Fasting-induced changes in the hypothalamic-pituitary-GH axis in the absence of GH expression: lessons from the spontaneous dwarf rat.

Fasting results in a reciprocal shift in hypothalamic neuropeptide Y (NPY) and GH-releasing hormone (GHRH) expression in the adult male rat. It is hypothesized that the fasting-induced rise in NPY is responsible for the GHRH decline and subsequent attenuation of pulsatile GH release. Fasting also leads to a decrease in circulating IGF-I, attributed to both reduced GH release and peripheral GH resistance. Although pituitary GH output is suppressed in the fasted rat, we report herein that pituitary GHRH receptor (GHRH-R) and GH secretagogue receptor (GHS-R) mRNA levels are increased, while pituitary expression of the somatostatin receptor subtype 2 (sst2) and 5 (sst5) is decreased, as determined by real-time reverse transcription (RT)-PCR. A shift in the expression of pituitary receptor subtypes to favor GH synthesis and release may be due, at least in part, to a decline in GH/IGF-I negative feedback. In order to test this hypothesis, we compared hypothalamic and pituitary response to fasting (72 h) in normal male rats and rats with isolated GH deficiency (spontaneous dwarf rats (SDR)). Circulating GH levels were undetectable in SDR, and IGF-I levels were less than 10% of normal controls. Fasting stimulated NPY mRNA levels in SDR; however, the rise in NPY mRNA levels was not accompanied by a fall in GHRH mRNA, as observed in fasted normal rats. In fact, GHRH mRNA levels paradoxically rose in the fasted SDR to 135% of fed controls. At the pituitary level, fasting did not alter sst2 and sst5 mRNA levels in SDR but did stimulate the expression of GHRH-R and GHS-R to 165% and 149% of fed controls, respectively. These results demonstrate that the fasting-induced changes in pituitary expression of sst2 and sst5, but not GHRH-R and GHS-R, are GH/IGF-I dependent. In addition, these results argue against the theory that the negative association of NPY and GHRH expression observed following fasting represents a simple cause-and-effect relationship and suggest that GH, either directly or indirectly, mediates the effects of fasting on hypothalamic GHRH expression.