RESUMO
Intracerebroventricular administration of human pancreatic growth hormone-releasing factor caused a dose-dependent inhibition of growth hormone secretion, elevated plasma glucose concentrations, and produced marked behavioral and motor effects. Immunoneutralization with antiserum to somatostatin did not reverse the suppression of growth hormone. These findings suggest that hypothalamic growth hormone-releasing factor may regulate its own neurosecretion through an "ultrashort-loop" negative feedback mechanism and may have important neurotransmitter and neuromodulatory functions in the brain.
Assuntos
Comportamento Animal/efeitos dos fármacos , Glicemia , Hormônio Liberador de Hormônio do Crescimento/farmacologia , Hormônio do Crescimento/metabolismo , Animais , Relação Dose-Resposta a Droga , Retroalimentação , Masculino , Ratos , Ratos EndogâmicosRESUMO
Intracerebroventricular administration of ILA's, a preparation enriched in insulin-like growth factors, caused a marked decrease in growth hormone secretory episodes and in body weight associated with reduced food intake over 24 hours. Central injection of insulin and bovine serum albumin had no such effects. These findings suggest that insulin-like growth factors play a role in growth hormone negative feedback and body weight regulation at the level of the central nervous system.
Assuntos
Peso Corporal/efeitos dos fármacos , Encéfalo/fisiologia , Hormônio do Crescimento/fisiologia , Insulina/farmacologia , Peptídeos/farmacologia , Somatomedinas/farmacologia , Animais , Encéfalo/efeitos dos fármacos , Ingestão de Alimentos/efeitos dos fármacos , Hormônio do Crescimento/antagonistas & inibidores , Hormônio do Crescimento/sangue , Insulina/sangue , Masculino , Ratos , Ratos EndogâmicosRESUMO
The mouse ob gene encodes leptin, an adipocyte hormone that regulates body weight and energy expenditure. Leptin has potent metabolic effects on fat and glucose metabolism. A mutation of the ob gene results in mice with severe hereditary obesity and diabetes that can be corrected by treatment with the hormone. In lean mice, leptin acutely increases glucose metabolism in an insulin-independent manner, which could account, at least in part, for some of the antidiabetic effect of the hormone. To investigate further the acute effect of leptin on glucose metabolism in insulin-resistant obese diabetic mice, leptin (40 ng x g(-1) x h(-1)) was administered intravenously for 6 h in C57Bl/6J ob/ob mice. Leptin increased glucose turnover and stimulated glucose uptake in brown adipose tissue (BAT), brain, and heart with no increase in heart rate. A slight increase in all splanchnic tissues was also noticed. Conversely, no increase in skeletal muscle or white adipose tissue (WAT) glucose uptake was observed. Plasma insulin concentration increased moderately but neither glucose, glucagon, thyroid hormones, growth hormone, nor IGF-1 levels were different from phosphate-buffered saline-infused C57Bl/6J ob/ob mice. In addition, leptin stimulated hepatic glucose production, which was associated with increased glucose-6-phosphatase activity. Conversely, PEPCK activity was rather diminished. Interestingly, hepatic insulin receptor substrate (IRS)1-associated phosphatidylinositol 3-kinase activity was slightly elevated, but neither the content of glucose transporter GLUT2 nor the phosphorylation state of the insulin receptor and IRS-1 were changed by acute leptin treatment. Hepatic lipid metabolism was not stimulated during the acute leptin infusion, since the content of triglycerides, glycerol, and citrate was unchanged. These findings suggest that in ob/ob mice, the antidiabetic antiobesity effect of leptin could be the result of a profound alteration of glucose metabolism in liver, BAT, heart, and consequently, glucose turnover. Insulin resistance of skeletal muscle and WAT, while not affected by acute leptin treatment, could also be corrected in the long term and account for some of leptin's antidiabetic effects.
Assuntos
Glucose/metabolismo , Músculo Esquelético/metabolismo , Proteínas/farmacologia , Animais , Eletrocardiografia , Feminino , Glucagon/sangue , Gluconeogênese , Hormônio do Crescimento/sangue , Frequência Cardíaca , Infusões Intravenosas , Insulina/sangue , Fator de Crescimento Insulin-Like I/metabolismo , Leptina , Fígado/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , Obesidade/metabolismo , Proteínas/administração & dosagem , Proteínas/metabolismoRESUMO
Recently, we have succeeded in using nonautologous myoblasts engineered to secrete mouse growth hormone (GH) to correct partially the growth retardation of the Snell dwarf mice, which suffer from pituitary GH deficiency. The allogeneic myoblasts were protected from immune rejection by enclosure in permselective microcapsules fabricated from alginate, thus validating the clinical efficacy of using universal nonautologous cells for somatic gene therapy. Because GH therapy is considered also for treating patients with normal pituitary function, we now apply this protocol to treat normal mice to evaluate the potential consequences of using GH gene therapy in subjects with no demonstrated GH deficiency. When microencapsulated allogeneic myoblasts engineered to secrete mouse GH were implanted into normal male and female mice, contrary to expectation, the treated animals became significantly shorter and lost weight; their internal organs became smaller and their tibial growth plates were less differentiated, indicating reduced skeletal growth. Females were more severely affected than males and 2 animals died by day 13 of unknown cause. By day 70, most of the abnormalities were restored to normal except for body weights, which remained below normal. In conclusion, although somatic gene therapy for GH delivery is beneficial for pituitary dwarfism, it may have adverse metabolic consequences in those with normal hypothalamic-pituitary functions, and the female mice were more severely affected than males.
Assuntos
Expressão Gênica , Hormônio do Crescimento/genética , Animais , Linhagem Celular , Transplante de Células , Composição de Medicamentos , Feminino , Terapia Genética/efeitos adversos , Crescimento , Transtornos do Crescimento/terapia , Hormônio do Crescimento/sangue , Hormônio do Crescimento/deficiência , Humanos , Fator de Crescimento Insulin-Like I/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Resultado do TratamentoRESUMO
Most of the currently approved human gene therapy protocols depend on genetic modification of autologous cells. We propose an alternate and potentially more cost-effective approach by implanting genetically modified "universal" cell lines to deliver desired gene products to nonautologous recipients. The recombinant allogeneic cells are protected from rejection after implantation by enclosure within immuno-protective alginate-poly-L-lysine-alginate microcapsules. The clinical efficacy of this strategy is now demonstrated by implanting microencapsulated allogeneic myoblasts engineered to secrete mouse growth hormone into the growth hormone-deficient Snell dwarf mice. The treated mutants attained increases in linear growth, body weights, peripheral organ weights, and tibial growth plate thickness significantly greater than those of the untreated controls. Secondary response to the exogenous growth hormone stimulation also resulted in increased fatty acid metabolism during the first month post-implantation. The microcapsules retrieved after about 6 months of implantation appeared intact. The encapsulated myoblasts retained a viability of > 60% and continued to secrete mouse growth hormone. Thus, implantation of nonautologous recombinant cells corrected partially the pleiomorphic effects of a transcription factor mutation in the Snell dwarf mice and the encapsulated cells remained functional for at least 6 months. This simple method of delivery recombinant gene products in vivo is a benign procedure, obviates the need for patient-specific genetic modification, and is amenable to industrial-scale quality control. It should have wide applications in therapies requiring a systemic continuous supply of recombinant gene products.
Assuntos
Transplante de Células/métodos , Sistemas de Liberação de Medicamentos/métodos , Transtornos do Crescimento/tratamento farmacológico , Hormônio do Crescimento/administração & dosagem , Proteínas Recombinantes/administração & dosagem , Alginatos , Animais , Materiais Biocompatíveis , Peso Corporal/efeitos dos fármacos , Desenvolvimento Ósseo/efeitos dos fármacos , Linhagem Celular , Composição de Medicamentos/métodos , Ácidos Graxos não Esterificados/sangue , Terapia Genética/métodos , Hormônio do Crescimento/sangue , Hormônio do Crescimento/deficiência , Hormônio do Crescimento/genética , Hormônio do Crescimento/farmacologia , Masculino , Camundongos , Músculos/citologia , Músculos/embriologia , Tamanho do Órgão/efeitos dos fármacos , Polilisina/análogos & derivados , Proteínas Recombinantes/farmacologia , TransfecçãoRESUMO
The ability of growth hormone (GH() to regulate its own secretion at the level of the central nervous system was examined in chronically cannulated freely-moving rats. Six-hour GH secretory profiles were obtained from 2 groups of rats administered either normal saline or rat GH via the lateral ventricle of the brain. The typical pulsatile pattern of GH secretion was evident in normal saline-treated control rats with most peak GH values > 400 ng/ml. Injection of rat GH (15 microgram/10 microliter) resulted in a significant suppression in amplitude of GH secretory pulses after an interval of 1 hour and plasma GH levels remained markedly depressed for up to 6 h after injection. Peak GH values did not exceed 76 ng/ml during the latter 3 h of the sampling period. These findings clearly demonstrate that GH can regulate its own secretion via a negative feedback system. Furthermore, the results suggest that the central nervous system is an important site of action for GH autoregulation.
Assuntos
Encéfalo/fisiologia , Retroalimentação , Hormônio do Crescimento/metabolismo , Animais , Encéfalo/efeitos dos fármacos , Hormônio do Crescimento/sangue , Hormônio do Crescimento/farmacologia , Cinética , Masculino , RatosRESUMO
A striking sexual dimorphism exists in the pattern of GH secretion and rate of somatic growth; however, the mechanism(s) mediating this sex difference is unknown. To elucidate the physiological roles of the hypothalamic neuropeptides, somatostatin (SRIF) and GRF, and their interrelation, in generating the sexually dimorphic GH secretory pattern we examined: 1) GH responsiveness to exogenous GRF and 2) the effects of immunoneutralization of endogenous SRIF and GRF on GH secretory dynamics, in free-moving male and female rats. In males, the GH response to 1 microgram rat(r)GRF(1-29)NH2 iv was significantly greater at peak compared to trough times of GH secretion (925.2 +/- 250.8 vs. 95.6 +/- 27.8 ng/ml; P less than 0.02), the latter known to be due to antagonization by the cyclic increased release of endogenous SRIF. In contrast, females failed to exhibit a time-dependent difference in GH responsiveness to GRF. Passive immunization with a specific antiserum to SRIF in males resulted in significant elevation of GH nadir levels but had no effect on GH peak amplitude. In contrast, immunoneutralization of endogenous SRIF in females caused a marked augmentation of plasma GH levels at all time points; there was a significant increase in GH peak amplitude (171.3 +/- 39.9 vs. 67.5 +/- 11.3 ng/ml; P less than 0.05), GH nadir (18.3 +/- 2.7 vs. 5.8 +/- 1.1 ng/ml; P less than 0.01) and mean 6-h plasma GH level (78.7 +/- 4.1 vs. 33.1 +/- 5.8 ng/ml; P less than 0.001), compared to normal sheep serum-treated controls. These results indicate that the pattern of hypothalamic SRIF secretion in females does not follow the male-like ultradian rhythm. Passive immunization with a specific antiserum to GRF obliterated spontaneous GH pulses in both sexes. Moreover, in females, anti-GRF serum attenuated GH nadir levels (4.3 +/- 1.7 vs. 21.4 +/- 3.5 ng/ml; P less than 0.01) indicating a physiological role for GRF in maintaining the elevated basal GH level of females, in addition to its important role in generating the episodic GH pulses. Taken together, these findings provide support for the hypothesis that, in female rats, the pattern of hypothalamic SRIF secretion into hypophyseal portal blood is continuous, rather than cyclical, as in the male; whereas in the case of GRF secretion, in addition to steady-state release which occurs at a higher level in females than males, there is also episodic GRF bursting which does not follow a specific rhythm, as in the male.(ABSTRACT TRUNCATED AT 400 WORDS)
Assuntos
Hormônio Liberador de Hormônio do Crescimento/fisiologia , Hormônio do Crescimento/metabolismo , Caracteres Sexuais , Transdução de Sinais , Somatostatina/fisiologia , Animais , Feminino , Hormônio Liberador de Hormônio do Crescimento/sangue , Hormônio Liberador de Hormônio do Crescimento/imunologia , Soros Imunes/imunologia , Imunização Passiva , Masculino , Fluxo Pulsátil , Ratos , Ratos Endogâmicos , Somatostatina/sangue , Somatostatina/imunologiaRESUMO
We examined the effects of intracerebroventricular (icv) administration of the somatostatin peptides, S-14 and S-28, and an analog, [D-Trp22]S-28, on spontaneous GH and glucose secretion in freely moving rats bearing chronic icv and intraatrial cannulae. Normal saline icv-treated control rats exhibited the typical pulsatile pattern of GH secretion. Central injection of S-14, S-28, and [D-Trp22]S-28 (each at two different doses, 5 and 10 micrograms) caused an early significant suppression of plasma GH levels which was of longer duration after S-28 or [D-Trp22]S-28 than after S-14 injection. The icv administration of S-28 and [D-Trp22]S-28 (but not S-14) also resulted in significant hyperglycemia, which persisted for periods up to 1 h. In a second study designed to determine whether somatostatin administered via the brain ventricle can reach the peripheral circulation, we measured plasma levels of S-14-like immunoreactivity (S-14 LI) and S-28-(15-28) LI at frequent time intervals after icv injection of S-14 (5 micrograms) and S-28 (10 micrograms). Plasma S-14 LI rose from a basal value of 0.16 +/- 0.02 (+/- SE) ng/ml to a peak of 3.0 +/- 1.0 ng/ml at 1 min, and S-28 icv resulted in a 150-fold increase in plasma S-28-(15-28) LI 1 min after injection. Sephadex G-50 gel chromatography revealed that the plasma immunoreactivity consisted of a single molecular species (either S-14 or S-28) corresponding to the centrally administered counterpart. These results demonstrate that after icv administration at high doses, both forms of somatostatin cause an acute inhibition of spontaneous GH secretion and that the larger form also causes hyperglycemia. We correlate these events with massive hypersomatostatinemia resulting from leakage of the somatostatin peptides from the cerebrospinal fluid into the systemic circulation. The findings indicate that icv injected hypothalamic hormones can reach the peripheral circulation and exert significant biological actions on distant target organs; thus, they have important implications for the design and interpretation of experiments in which peptides are administered via the cerebral ventricles.
Assuntos
Glicemia/metabolismo , Hormônio do Crescimento/metabolismo , Somatostatina/sangue , Animais , Cromatografia em Gel , Injeções Intraventriculares , Masculino , Ratos , Ratos Endogâmicos , Somatostatina/administração & dosagem , Somatostatina/análogos & derivados , Somatostatina/farmacologia , Somatostatina-28RESUMO
Restriction of dietary protein stunts growth in the rat, but the mechanism is not well understood. In the present study, we examined the effects of dietary protein restriction on spontaneous and GH-releasing factor (GRF)-stimulated GH release and assessed the possible involvement of endogenous somatostatin (SRIF). Spontaneous 6-h plasma GH profiles were obtained from free-moving adult male rats fed either a 23% (normal) or 4% (low) isocaloric protein diet. Control rats exhibited the typical pulsatile pattern of GH release. In contrast, rats fed the low protein diet showed a significant reduction in GH peak amplitude (85.0 +/- 10.4 vs. 171.3 +/- 20.5 ng/ml; P < 0.01) and mean 6-h plasma GH level (18.1 +/- 2.0 vs. 40.9 +/- 6.0 ng/ml; P < 0.01) as early as 4 days after diet onset and a more than 3-fold suppression of GH pulse amplitude by 7 days. Although protein-restricted animals exhibited the typical cyclic responsiveness to 1 microgram rGRF-(1-29)NH2 i.v., the magnitude of the GH response to GRF challenge was attenuated 3- to 4-fold in these rats compared to that in normal diet-fed controls. Passive immunization of protein-restricted rats with SRIF antiserum resulted in a significant augmentation of both GH pulse amplitude (115.3 +/- 16.7 vs. 36.0 +/- 2.8 ng/ml; P < 0.01) and mean 6-h plasma GH level (34.4 +/- 5.0 vs. 10.0 +/- 1.6 ng/ml; P < 0.01) compared to those in protein-deprived rats administered normal sheep serum. Pituitary size (7.8 +/- 0.2 vs. 12.1 +/- 0.4 mg; P < 0.001) and pituitary GH content (320.5 +/- 18.9 vs. 526.6 +/- 26.8 micrograms; P < 0.001) were markedly reduced after 3-week maintenance on the 4% protein diet. In a separate study, rats fed 70% of the control diet (calorically equivalent to that consumed by rats fed 4% protein) showed no significant alteration in pulsatile GH release, thus excluding caloric restriction as a cause of the GH suppression. These results demonstrate that lack of dietary protein 1) blunts spontaneous pulsatile GH release, 2) attenuates GH responsiveness to GRF challenge, and 3) reduces pituitary GH content and size. Our findings suggest that the low protein-induced suppression of GH release is mediated at least in part by increased SRIF secretion. Such impairments in the GH neuroendocrine axis probably contribute to the growth retardation observed in this model.
Assuntos
Proteínas Alimentares/administração & dosagem , Hormônio Liberador de Hormônio do Crescimento/farmacologia , Hormônio do Crescimento/metabolismo , Deficiência de Proteína/fisiopatologia , Animais , Ingestão de Energia , Privação de Alimentos , Imunização Passiva , Masculino , Periodicidade , Ratos , Ratos Sprague-Dawley , Proteínas Recombinantes/farmacologia , Somatostatina/imunologia , Somatostatina/fisiologiaRESUMO
Insulin-like growth factor (IGF)-I and -II peptides, receptors, mRNAs, and binding proteins are widely distributed in the central nervous system (CNS), yet their physiological role in the brain remains largely unknown. While earlier in vivo studies in the rat suggested that IGF-I may participate in feedback regulation of GH secretion at a CNS level, the preparations used were only partially pure. The recent availability of purified recombinant IGF-I and -II peptides prompted us to reexamine the involvement of the IGFs in vivo in central regulation of pulsatile GH secretion. Five groups of free-moving adult male rats bearing chronic intracerebroventricular (icv) and intracardiac venous cannulae were icv administered IGF-I (in doses of 0.5, 2, 3, and 10 micrograms) or the acid-saline vehicle; an additional group received 1 microgram of the potent IGF-I analog, long R3 IGF-I. Spontaneous 6-h plasma GH secretory profiles were obtained from all groups. Vehicle-injected control animals exhibited the typical pulsatile pattern of GH secretion, with most peak GH values above 150 ng/ml and trough levels below 1.2 ng/ml. Central administration of IGF-I alone or long R3 IGF-I at all doses tested failed to alter the pulsatile pattern of GH release; there were no significant differences in GH peak amplitude, GH trough level, GH interpeak interval, or mean 6-h plasma GH level compared to those in vehicle-injected controls. In a second study, designed to determine the effects of central administration of IGF-I and IGF-II, in combination, icv injection of 1 microgram IGF-I and 1 microgram IGF-II resulted in a marked suppression in the amplitude of spontaneous GH secretory bursts approximately 3 h after injection; both GH pulse amplitude (43.5 +/- 5.6 vs. 130.6 +/- 14.6 ng/ml; P less than 0.001) and mean plasma GH level (16.3 +/- 1.9 vs. 35.2 +/- 1.8 ng/ml; P less than 0.001) were severely reduced 3-6 h after injection compared to those in vehicle-injected controls. These results demonstrate that IGF-I alone does not play a physiologically important role in feedback regulation of GH secretion at the level of the CNS. Our findings suggest a synergistic interaction between IGF-I and -II in the brain for central control of pulsatile GH secretion.
Assuntos
Hormônio do Crescimento/metabolismo , Fator de Crescimento Insulin-Like II/farmacologia , Fator de Crescimento Insulin-Like I/farmacologia , Periodicidade , Animais , Glicemia/metabolismo , Sinergismo Farmacológico , Injeções Intraventriculares , Insulina/sangue , Fator de Crescimento Insulin-Like I/administração & dosagem , Fator de Crescimento Insulin-Like II/administração & dosagem , Masculino , Ratos , Ratos Endogâmicos , Proteínas Recombinantes/farmacologiaRESUMO
Calcitonin (CT) binds to specific receptors in the hypothalamus and has been localized in the pituitary, suggesting a potential neuroendocrine role for this peptide. We and others have previously shown that CT given centrally markedly suppresses pulsatile GH secretion. However, the mechanism mediating this response remains to be elucidated. In the present study, we assessed the involvement of the two hypothalamic GH-regulatory peptides, somatostatin (SRIF) and GH-releasing factor (GRF), using a combination of in vivo and in vitro techniques. Six-hour GH secretory profiles were obtained from eight groups of freely moving rats bearing chronic intracerebroventricular (icv) and intraatrial cannulae. In four groups, salmon (s) CT (250 ng/10 microliters) was administered icv, whereas the remaining four groups received either normal saline (NS) icv or sCT iv. Central injection of sCT caused a severe suppression in amplitude of spontaneous GH pulses compared to NS icv-treated control rats, whereas the same dose of sCT iv had no significant effect. Passive immunization of sCT icv-injected rats with a specific antiserum to SRIF failed to restore the amplitude of GH pulses to normal values. In addition, in vitro basal and 50 mM K+-stimulated SRIF release from incubated hypothalamic fragments was not altered by sCT in doses ranging from 10(-10) to 10(-6) M. The iv administration of a bolus of rat GRF (1-29)NH2 (1 microgram) 1 h after sCT icv injection also failed to augment plasma GH levels compared to sCT iv-treated rats (16.6 +/- 10.0 vs. 326.6 +/- 63.6 ng/ml; P less than 0.001) and NS icv controls (407.2 +/- 145.4 ng/ml; P less than 0.01). Blood calcium levels decreased similarly 1 h after iv and icv sCT administration. These results demonstrate that: sCT inhibits pulsatile GH secretion via a central nervous system site of action, GH suppression induced by sCT is apparently not due solely to increased hypothalamic SRIF release, and centrally administered sCT produces an acute loss of responsiveness of somatotrophs to GRF, which can be dissociated from peripheral blood calcium levels.
Assuntos
Calcitonina/farmacologia , Hormônio Liberador de Hormônio do Crescimento/fisiologia , Hormônio do Crescimento/metabolismo , Somatostatina/fisiologia , Animais , Cálcio/sangue , Hormônio do Crescimento/antagonistas & inibidores , Hormônio Liberador de Hormônio do Crescimento/farmacologia , Hipotálamo/metabolismo , Soros Imunes/farmacologia , Masculino , Hipófise/metabolismo , Ratos , Ratos Endogâmicos , Somatostatina/imunologiaRESUMO
Exogenous GH is known to exert a negative feedback effect on its own responsiveness to GH-releasing factor (GRF); however, the mechanism is not known. In the present study we examined the time course of effects of a single sc administration of recombinant human (rh) GH on GH responsiveness to GRF and investigated the possible involvement of somatostatin (SRIF) in this response. Free-moving adult male rats were administered 200 micrograms rhGH, sc, at 0800 h and subsequently challenged with 1 microgram GRF-(1-29)NH2, iv, at times of spontaneous peaks (1100 and 1500 h) and troughs (1300 h) in GH secretion during a 6-h (1000-1600 h) sampling period. H2O-injected control rats exhibited the typical cyclic responsiveness to GRF stimulation, with GRF-induced GH release significantly greater during peak compared to trough periods of the GH rhythm. Pretreatment with rhGH 3 h before GRF injection markedly inhibited the GH response to GRF at a peak time [integrated GH release over 30 min, 1135 +/- 271 vs. 6372 +/- 1185 ng/ml.30 min in H2O-injected controls (mean +/- SE); P less than 0.01]. In striking contrast, 5 h after rhGH administration, there was a 6-fold augmentation of GH responsiveness to GRF compared to that in H2O-injected controls at a trough time (7032 +/- 1622 vs. 1128 +/- 216 ng/ml.30 min; P less than 0.01). High GH responsiveness to GRF was preserved 7 h after rhGH injection. Passive immunization of rhGH-treated rats with SRIF antiserum reversed the rhGH-induced blunted GH response at 3 h (7985 +/- 366 vs. 1705 +/- 431 ng/ml.30 min in rhGH-treated control rats given normal sheep serum; P less than 0.01) and completely restored GH responsiveness to levels as high as those in H2O-injected controls. These results demonstrate that 1) a single sc injection of rhGH markedly attenuates GH responsiveness to GRF acutely for about 3 h, but subsequently enhances somatotroph sensitivity to the stimulatory actions of GRF; and 2) the short term blunting of GRF-induced GH release by rhGH is due at least in part to increased release of endogenous SRIF. The subsequent potentiation of GH responsiveness to GRF is probably due to a SRIF-mediated build-up of pituitary GH stores in a readily releasable pool. Such a mechanism of GH autofeedback may play a physiological role in the genesis of pulsatile GH secretion.
Assuntos
Hormônio Liberador de Hormônio do Crescimento/farmacologia , Hormônio do Crescimento/sangue , Somatostatina/fisiologia , Animais , Retroalimentação , Hormônio do Crescimento/farmacologia , Soros Imunes/imunologia , Imunização Passiva , Masculino , Ratos , Ratos Endogâmicos , Proteínas Recombinantes/farmacologia , Somatostatina/imunologia , Fatores de TempoRESUMO
Endogenous pulsatile GH secretion is blunted by the administration of exogenous GH; however, few data are available on the time course of GH negative feedback, and the mechanism by which this occurs still remains unclear. In the present study, we examined the temporal pattern of the inhibitory effect induced by an acute (single) and chronic (5 days) sc recombinant human (rh) GH injection regimen on spontaneous GH release in the rat and assessed the possible involvement of the hypothalamic GH-inhibitory peptide, somatostatin (SRIF), in this response. Eight-hour (0800-1600 h) GH secretory profiles, obtained from free-moving adult male rats administered a single sc injection of 200 micrograms rhGH at 0800 h, revealed a marked suppression of spontaneous GH pulses (GH peak amplitude: 45.7 +/- 10.9 vs. 207.8 +/- 31.7 ng/ml in H2O-injected control rats; P less than 0.001) lasting for up to 4.1 +/- 0.1 h after the injection (mean 4-h plasma GH level: 13.6 +/- 3.6 vs. 49.4 +/- 7.0 ng/ml in H2O-injected controls; P less than 0.01). During the subsequent 4- to 8-h period, recovery of spontaneous GH secretory bursts was evident, and neither the GH peak amplitude nor mean 4-h plasma GH level of rhGH-treated rats was significantly different from that of H2O-injected controls. The magnitude, time course, and recovery of the rhGH-induced inhibitory effect on pulsatile GH release after chronic rhGH treatment was similar to that after a single injection. Passive immunization of rhGH-treated rats with SRIF antiserum reversed the rhGH-induced inhibition of spontaneous GH pulses (peak amplitude: 131.7 +/- 53.7 vs. 7.1 +/- 3.4 ng/ml in rhGH-treated control rats given normal sheep serum; P less than 0.05) and restored both the GH peak amplitude and mean plasma GH level to values similar to those in H2O-injected controls. Taken together, these results demonstrate that: 1) the inhibitory effect of rhGH on endogenous pulsatile GH release is of short duration (approximately 4 h); 2) the time course of this response does not change after 5-day repeated rhGH administration; and 3) the feedback effect of GH on its own spontaneous release is exerted, at least in part, by increasing hypothalamic SRIF secretion. Such a mechanism of GH feedback may be important in the physiological control of pulsatile GH secretion.
Assuntos
Hormônio do Crescimento/metabolismo , Hormônio do Crescimento/farmacologia , Somatostatina/fisiologia , Ciclos de Atividade , Animais , Esquema de Medicação , Retroalimentação , Soros Imunes , Imunização Passiva , Fator de Crescimento Insulin-Like I/metabolismo , Masculino , Ratos , Ratos Endogâmicos , Proteínas Recombinantes/farmacologia , Somatostatina/imunologiaRESUMO
Sequential blood samples were obtained from undisturbed freely-behaving male rats bearing chronic intracardiac venous cannulae. Blood was withdrawn every 15 min for periods of 4-24 h; plasma was separated, and saline-resuspended red cells were reinjected. Plasma GH was determined by radioimmunoassay. Pulsatile GH secretion was evident in each animal with most peak values greater than 200 ng/ml and most trough values less than ng/ml. The GH secretory episodes occurred at approximately 3 h intervals, and this rhythmic pattern of GH secretion persisted unchanged across all phases of a 12-h light-dark (L-D) cycle. Seven major episodes of GH secretion were observed during a single 24-h period. The mean period, or time interval between episodes, in 24 animals was 3.32 +/- 0.07 (SEM)h. The timing of the pulses with respect to the L-D cycle was similar in most animals, indicating that the rhythm may be entrained to the L-D cycle. The role of environmental lighting was further assessed in 14 animals exposed to constant light for 7 weeks. The results show that the basic rhythm was unchanged (mean period 3.18 +/- 0.06 h, peaks greater than 200 ng/ml, troughs less than 1 ng/ml), although entrainment to time of day was not evident. Subsequent exposure to the 12-h L-D cycle resulted in reversion to an entrained rhythm. These results suggest 1) that GH secretion in the rat is governed by an endogenous ultradian rhythm, with a periodicity of approximately 3.3 h, and 2) that the alternation of light and darkness probably serves as a Zeitgeber which sets the biological "clock" for GH secretion, but is not necessary for maintenance of the basic rhythm.
Assuntos
Relógios Biológicos , Ritmo Circadiano , Hormônio do Crescimento/metabolismo , Ratos/fisiologia , Animais , Escuridão , Hipotálamo/fisiologia , Luz , MasculinoRESUMO
We have examined the capacity of calcitonin gene-related peptide (CGRP) to exert two biological actions in the central nervous system (CNS): modulation of GH release and alteration of food intake. These effects were compared with those of calcitonin (CT). Intracerebroventricular administration of both rat (r) CGRP (10 micrograms/10 microliter) and salmon (s) CT (250 ng/10 microliter) to chronically cannulated freely-moving rats reduced the frequency and amplitude of spontaneous GH secretory pulses; however, the suppressive effect was of longer duration with sCT than with rCGRP. Both peptides also significantly reduced spontaneous food intake over a 6-h period when administered centrally, although the decrease with sCT was again more extensive than that with rCGRP. The results demonstrate that CGRP can simulate two of the effects of CT in brain. Furthermore, the findings suggest that CGRP may function as an endogenous ligand for CT receptors in the CNS, and participate centrally in the modulation of GH release and the control of satiety.
Assuntos
Encéfalo/efeitos dos fármacos , Calcitonina/farmacologia , Ingestão de Alimentos/efeitos dos fármacos , Hormônio do Crescimento/metabolismo , Proteínas do Tecido Nervoso/farmacologia , Animais , Peptídeo Relacionado com Gene de Calcitonina , Masculino , Ratos , Ratos Endogâmicos , Receptores da Calcitonina , Receptores de Superfície Celular/efeitos dos fármacosRESUMO
We examined the effects of 2-deoxy-D-glucose (2DG)-induced intracellular glucoprivation on GH, insulin, and glucose secretory dynamics in freely moving rats bearing chronic intracerebroventricular and intracardiac venous cannulae. Intravenous administration of 2DG (400 mg/kg) caused a severe suppression in amplitude and duration of spontaneous GH surges; plasma GH levels remained significantly depressed for at least 5 h in the presence of marked hyperglycemia. Plasma insulin levels were unchanged. Central administration of a low dose of 2DG (8 mg/10 microliters) also markedly attenuated GH pulse amplitude and raised plasma glucose levels, but this low dose was without effect when injected peripherally, suggesting a central site of action. To elucidate the mechanism(s) mediating the GH suppression response to insufficient glucose we assessed the involvement of the two hypothalamic GH-regulatory peptides, somatostatin (SRIF) and GH-releasing factor (GRF). Passive immunization of 2DG-treated rats with a specific SRIF antiserum (AS) caused an initial surge of GH release and significant elevation of both trough and mean 6-h plasma GH levels, compared to 2DG-normal sheep serum controls. However, SRIF AS failed to restore the amplitude of GH pulses to normal levels. Administration of three iv boluses of human GRF (10 micrograms), at 90-min intervals, to 2DG-treated rats resulted in GH release which was variable and time dependent; the magnitude of the first response (1 h after 2DG injection) was significantly less than that of the other two. Immunoneutralization with SRIF AS eliminated this difference and significantly enhanced human GRF-induced GH release. These results demonstrate that intracellular glucopenia is a potent inhibitor of pulsatile GH secretion in the rat and that this response is mediated, in part, by an increase in SRIF release. While the present findings are compatible with the hypothesis that glucoprivation induces an acute SRIF release, only a partial role for SRIF is indicated in this response. The longer lasting suppression of GH pulses observed after glucose deprivation may be due to decreased output of GRF from the hypothalamus.
Assuntos
Desoxiaçúcares/farmacologia , Desoxiglucose/farmacologia , Hormônio do Crescimento/sangue , Somatostatina/farmacologia , Animais , Glicemia/metabolismo , Hormônio Liberador de Hormônio do Crescimento/fisiologia , Hipotálamo/efeitos dos fármacos , Imunização Passiva , Injeções Intraventriculares , Insulina/sangue , Masculino , Ratos , Ratos Endogâmicos , Somatostatina/fisiologiaRESUMO
To further delineate the relationship between GH-releasing factor (GRF) and somatostatin (SRIF) in generation of the ultradian rhythm of GH secretion, we used two GRF peptides, human pancreas (hp) GRF-44 and rat hypothalamic (rh) GRF, and studied their interaction with SRIF by passive immunization with a specific antiserum (AS) to SRIF. Freely moving, chronically cannulated male rats were given 10 micrograms of either hpGRF-44 or rhGRF, iv, during peak (1100 h) and trough (1300 h) periods of the GH rhythm. Six-hour plasma GH profiles were obtained after pretreatment with either SRIF AS or normal sheep serum (NSS) as a control. In NSS-treated rats, the plasma GH responses to both hpGRF-44 and rhGRF were significantly greater when the peptides were administered during peak than during trough periods. Immunoneutralization with SRIF AS eliminated these differences and permitted marked GH release in response to both peptides at 1300 h. In addition, SRIF AS augmented the GRF-induced GH response at 1100 h compared with that in NSS controls. The rhGRF peptide caused significantly more GH release than hpGRF under both conditions. These results demonstrate that 1) the GH-releasing abilities of the GRF peptides vary markedly according to the time of injection; 2) the weak GRF-induced GH response observed during trough periods of the GH rhythm is due to antagonization by endogenous circulating SRIF; and 3) the rat-derived GRF may be a more potent GH secretagogue than the human-derived peptide in the rat. The findings reported here together with the available evidence provide support for the hypothesis that GRF and SRIF are secreted tonically from the hypothalamus into the hypophyseal portal blood, and that superimposed upon this steady state release is an additional 3- to 4-h rhythmic surge of each peptide, providing for integration of the ultradian rhythm of GH secretion, as observed in peripheral blood.
Assuntos
Hormônio Liberador de Hormônio do Crescimento/fisiologia , Hormônio do Crescimento/metabolismo , Periodicidade , Somatostatina/fisiologia , Animais , Complexo Antígeno-Anticorpo , Hormônio do Crescimento/sangue , Humanos , Hipotálamo/fisiologia , Soros Imunes , Imunização Passiva , Masculino , Pâncreas/fisiologia , Ratos , Ratos Endogâmicos , Fatores de TempoRESUMO
Temporal patterns of plasma GH, immunoreactive insulin (IRI), and glucose were defined by obtaining serial blood samples from freely-moving male rats bearing chronic intracardiac venous cannulae. Blood was withdrawn every 15 min for periods of 6 h. Plasma GH and IRI were determined by radioimmunoassay. The typical ultradian rhythm of GH secretion was evident in each undisturbed animal (peaks greater than 200 ng/ml; troughs less than 1 ng/ml; mean period: 3.40 +/-0.08 h). Basal plasma IRI and glucose levels fluctuated minimally. There was no significant correlation between plasma GH and IRI, GH and glucose, or IRI and glucose levels in unfed rats. The rhythmic GH secretory patterns of feeding animals (mean period: 3.12 +/-0.16 h; peaks greater than 200 ng/ml; troughs less than 1 ng/ml) were similar to those of non-feeding animals (mean period: 3.34 +/-0.15 h; peaks greater than 200 ng/ml; troughs less than 1 ng/ml) despite large fluctuations in plasma IRI levels and a wide variation in the number and size of the meals taken. No consistent relation was observed between the ingestion of meals and the bursts of GH secretion. The mean period of the GH rhythm was not significantly altered by hyperglycemia (mean period; 3.25 +/- 0.08 h), although the amplitude of the pulses of half of the hyperglycemic rats was markedly depressed. Insulin-induced hypoglycemia caused a significant depression in the amplitude of the GH pulses; however, the pattern of this response was not consistent. Despite wide variability in the GH response, the magnitude and time course of recovery of the plasma glucose levels was similar in all animals. These results suggest that GH secretion in the rat is regulated primarily by an endogenous ultradian rhythm which is not dependent on changes in plasma glucose or IRI levels, and continues to function independently of feeding behavior. It is unlikely that GH is an important physiologic regulator of glucose homeostasis in this species.
Assuntos
Ritmo Circadiano , Hormônio do Crescimento/sangue , Hiperglicemia/sangue , Hipoglicemia/sangue , Insulina/sangue , Animais , Glicemia/metabolismo , Jejum , Hipoglicemia/induzido quimicamente , Luz , Masculino , RatosRESUMO
The ultradian rhythm of GH secretion in the male rat is generated by the reciprocal cyclic release of somatostatin (SRIF) and GH-releasing factor (GRF) from the hypothalamus. We recently demonstrated the presence of high affinity [125I]SRIF-binding sites on a subpopulation of GRF-containing arcuate neurons in rat hypothalamus. In the present study, we tested the hypothesis that these binding sites undergo rhythmic fluctuations in parallel with those of GH. Adult male rats were killed at times associated with either a peak (1100 h) or trough (1300 h) period of the GH rhythm. The hypothalamus was serially sectioned from the retrochiasmatic nucleus, rostrally, to the mammillary recess of the third ventricle, caudally, and [125I]SRIF-binding sites were labeled in vitro using high resolution autoradiography. [125I]SRIF-labeled neuronal perikarya were counted at eight cross-sectional levels across the arcuate nucleus, and binding densities were quantitated over each of these cross-sectional surfaces using computer-assisted microdensitometry. Both the number of labeled cells and the density of [125I]SRIF binding varied as a function of the time of death. The average number of [125I]SRIF-labeled cells per 10-microns thick section was 2- to 3-fold higher in rats killed at 1100 h than in those killed at 1300 h. In addition, overall binding density levels were 65% higher in animals killed at the onset of a GH peak than in those killed at the time of a GH trough. Both of these effects were apparent throughout the rostrocaudal extent of the arcuate nucleus. In contrast, [125I]SRIF binding density in the cerebral cortex did not vary between 1100 and 1300 h. These results demonstrate an ultradian variation in SRIF binding to arcuate neurons in relation to the peaks and troughs of the GH rhythm. Such rhythmicity in SRIF receptors might underlie a temporal responsiveness of arcuate GRF neurons to endogenous SRIF and may be an important mechanism by which SRIF modulates the rhythmic release of hypothalamic GRF in generation of the ultradian rhythm of GH secretion.
Assuntos
Ciclos de Atividade/fisiologia , Núcleo Arqueado do Hipotálamo/metabolismo , Somatostatina/metabolismo , Animais , Autorradiografia , Sítios de Ligação , Hormônio do Crescimento/metabolismo , Radioisótopos do Iodo , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
GH circulates in the plasma partially bound with a GH-binding protein (GHBP), but the physiological significance of the GHBP and how it affects GH bioactivity in vivo is still unknown. In the present study, we took advantage of the known biological action of exogenous human (h) GH to inhibit endogenous rat (r) pulsatile GH release and examined the effect of combining hGH with recombinant hGHBP on this response in normal rats. Spontaneous 7-h plasma rGH and hGH profiles were obtained from four groups of free-moving adult male rats s.c. administered either: 1) 200 microg hGH alone; 2) a mixture of 200 microg hGH and 200 microg hGHBP preincubated for 30 min before injection; 3) 200 microg hGHBP alone; or 4) Tris buffer (vehicle) alone. Rats administered the vehicle or hGHBP separately exhibited the typical pulsatile pattern of rGH secretion. Injection of hGH alone resulted in a marked (P < 0.01) suppression of spontaneous rGH pulses for approximately 3.5 h after the injection compared with vehicle-injected controls; during the subsequent 3.5- to 7-h period, recovery of spontaneous rGH peaks was evident. Plasma levels of hGH in these animals reached a peak within 1 h after hGH injection and declined to near undetectable levels by the end of the sampling period. In contrast, the disappearance rate of hGH was markedly slower in rats administered the hGH + hGHBP complex; plasma hGH concentrations at 7 h after injection were 14-fold higher than those in animals administered hGH alone, and hGH was still readily detectable up to 24 h after injection. However, despite the markedly higher levels of hGH persisting throughout the sampling period in complex-injected rats, both the time course of hGH-induced inhibition of rGH and the recovery of spontaneous rGH pulses were similar to those of animals administered hGH alone. Moreover, there were no significant modifications of plasma insulin-like growth factor-1 levels for up to 24 h after injection of the hGH + hGHBP complex. Computer simulations revealed that most of the total hGH observed during the 3.5- to 7-h period was circulating in the bound form. These results demonstrate that, despite hGHBP's ability to markedly prolong the bioavailability of hGH, precomplexing hGH with hGHBP failed to enhance hGH's in vivo bioactivity in the inhibition of endogenous pulsatile rGH release. Our findings do not provide support for the concept that the GHBP enhances the bioactivity of GH in vivo, at least over the time course examined here.