Mechanisms underlying attenuated contractile response of aortic rings to noradrenaline in fructose-fed mice.

We hypothesized that an impairment of endothelial dysfunction and an increased response to alpha-adrenoceptor agonists may occur in fructose-fed, insulin-resistant mice. The aim of the present study was to assess the relationship between endothelial dysfunction and agonist-induced contractile responses in such mice. The acetylcholine-induced relaxation was significantly attenuated in streptozotocin-diabetic and fructose-fed mice. The contractile response to noradrenaline was significantly weaker than the control in fructose-fed but not in streptozotocin-diabetic mice; treatment with N(G)-nitro-L-arginine effectively restored this response. Incubating aortic rings with noradrenaline increased the NO(x) [nitrite (NO(2)(-)) and nitrate (NO(3)(-))] level and this level was significantly higher in fructose-fed mice than in control mice. Clonidine induced a dose-dependent relaxation in aortic rings pre-contracted with prostaglandin F(2alpha) that was completely abolished by N(G)-nitro-L-arginine; this relaxation was markedly enhanced in fructose-fed mice. In both control and fructose-fed mice, the clonidine-induced relaxation was significantly attenuated and the noradrenaline-induced contraction augmented by pertussis toxin. These results suggest that endothelial function is attenuated in both fructose-fed and streptozotocin-diabetic mice. It is suggested that the decreased noradrenaline contractile response in fructose-fed mice (compared to both controls and streptozotocin-diabetic mice) may be due to an increase in nitric oxide formation mediated by endothelial GTP-binding-coupled alpha(2)-adrenoceptors.

Contractile responses in spontaneously diabetic mice. I. Involvement of superoxide anion in enhanced contractile response of aorta to norepinephrine in C57BL/KsJ(db/db) mice.

This study investigated the influence of superoxide anion on the norepinephrine (NE)-induced contractile response in spontaneously diabetic mice. In aortic rings with intact endothelium, NE elicited only a slight increase in tension in nondiabetic mice (db/+M), but a much greater dose-dependent contraction in spontaneously diabetic mice (db/db mice). The NE-induced contractile response was significantly reduced by pretreatment with SOD (180 U/ml) in diabetic mice, but not in control mice. The NE-induced contraction was significantly enhanced by pretreatment with diethyldithiocarbamic acid (DETCA, 10(-3) M), a Cu/Zn SOD inhibitor, in control mice, but not in diabetic mice. The dose-response curve for the acetylcholine-induced relaxation was slightly, but significantly attenuated in diabetic mice. When aortic rings from control mice were incubated with a mixture of hypoxanthine (10(-5) M), xanthine oxidase (0.1 U/ml) and catalase (1000 U/ml) in control mice, they gradually contracted. This contraction was abolished by pretreatment with SOD (180 U/ml) or indomethacin (10(-5) M) or by removal of the endothelium. The enhanced NE-induced dose-dependent contraction seen in diabetic mice was markedly attenuated by indomethacin. These results suggest that in db/db diabetic mice, superoxide anion, perhaps via vasoconstrictor prostanoids, may enhance the contraction induced by NE.

Contractile responses in spontaneously diabetic mice. II. Effect of cholestyramine on enhanced contractile response of aorta to norepinephrine in C57BL/KsJ (db/db) mice.

To examine the possible role of low-density lipoprotein (LDL) cholesterol in the enhanced norepinephrine (NE)-induced contractile response seen in spontaneously diabetic mice (db/db mice), we examined the effect of chronic administration of cholestyramine on the NE-induced contraction. Although chronic cholestyramine (300 mg/kg, po for 1 month) significantly lowered total cholesterol, high-density lipoprotein (HDL) cholesterol, LDL cholesterol, and triglyceride, the plasma glucose and insulin levels were unaffected. The enhanced NE response in diabetic mice was not affected by the chronic administration of cholestyramine. The K(+)-induced contractile response was not different among nondiabetic, diabetic, and diabetic mice chronically treated with cholestyramine. These results suggest that neither LDL cholesterol nor triglyceride is involved in the enhancement of the NE-induced contractile response seen in spontaneously diabetic mice.

[Pure shearing injury--clinical signs and symptoms, diagnosis, prognosis and review of the literature].

In some of the brains of head injury, shearing of axons and vessels caused by direct impact is seen widely in the white matter. This type of injury is designated by neuropathologists as shear injury or diffuse damage of impact type. Macroscopic lesions adjacent to the superior cerebeller peduncle and lesions of the corpus callosum are commonly seen in the brains with severe shearing injuries and the former is detected by CT scan as a high density at the quadrigeminal cistern, and this indicates occurrence of a severe shearing injury. Among 600 patients with head injuries, we managed 23 cases with pure shearing injury. These cases are classified into three groups according to the duration of coma. In the fulminant type (7 cases) the patients had severe brainstem signs including impairment of vital signs and died immediately after the injury. In the severe type (8 cases) the patients were decerebrated with normal pupillary response on admission and were severely disabled (i.e. tremor, truncal ataxia, dementia). In the moderate type (8 cases), the patients had only disturbance of consciousness for a long period, and recovered almost to the pre-injury state. Follow-up CT scan showed enlargement of the ventricles and sulci.

Long-term growth hormone responses to nonspecific hypothalamic hormones in acromegalic patients.

Nonspecific hypothalamic hormones such as thyrotropin-releasing hormone or luteinizing hormone-releasing hormone, or both, elicited abnormal growth hormone responses in 73 of 108 (67.6%) acromegalic patients. After transsphenoidal adenomectomy, the provocative tests using these hormones were repeated in 26 patients with abnormal preoperative growth hormone responses to study variations in these responses during a 1-8-year observation period (average duration, 4 years). After surgery, 7 of the 26 patients regained normal basal growth hormone levels (less than 5 ng/mL) and manifested normal responses to the hypothalamic hormones. During the postoperative observation period, their basal growth hormone levels remained normal as did their responses to provocation with hypothalamic hormones, confirming that the adenoma had been completely resected. Eight other patients demonstrated normal growth hormone levels after surgery; however, they continued to have abnormal responses to provocation with hypothalamic hormones, suggesting the presence of residual adenomatous tissue in the gland. These patients manifested no marked increase in basal or peak growth hormone levels during the follow-up period (from less than 1 to less than 7.5 years) and they were all in clinical remission without any other treatment. Only one incompletely adenomectomized patient who had received no additional treatment experienced regrowth of the tumor. The main factor affecting the surgical results appears to be the preoperative basal growth hormone level, because abnormal growth hormone secretion ceased in all patients who had manifested preoperative levels below 45 ng/mL. Technical refinements of the operative procedure are another important factor in the postoperative outcome. Peritumoral tissue resection after simple selective adenomectomy is mandatory for better surgical results. Our studies indicate that fairly good results can be obtained without risk of the recurrence of the tumor or regrowth, when postoperative growth hormone levels are below 5 ng/mL and that the results are not affected by the postoperative growth hormone responses to provocation with hypothalamic hormones.

Levels of human and rat hypothalamic growth hormone-releasing factor as determined by specific radioimmunoassay systems.

Polyclonal antibodies to synthetic human pancreatic growth hormone-releasing factor [hpGRF(1-44)NH2] and rat hypothalamic growth hormone-releasing factor [rhGRF(1-43)OH] were produced in rabbits by injecting these weak immunogens, coupled to thyroglobulin and emulsified with complete Freund's adjuvant in the presence of activated charcoal, directly into the spleen. A subsequent booster injection by the conventional intramuscular route resulted in high-titer antibodies, which at a 1:20,000 dilution were used to develop highly sensitive and specific radioimmunoassays for these peptides. By using antibodies with an apparent Ka of 3.3 X 10(-12) (human) and 7.7 X 10(-11) (rat), the sensitivity of these assays in both human and rat was found to be less than 1 fmol. The antibody to hpGRF(1-44)NH2 is directed against the COOH-terminal region of the molecule, as shown by its crossreactivity with various hpGRF analogues: 140% with hpGRF(30-44)NH2; 1%-2% with hpGRF(1-37)OH, hpGRF(1-40)OH, and hpGRF(1-40)NH2; and none with hpGRF(1-29)NH2. Serial dilutions of human and rat hypothalamic extracts demonstrated parallelism with the corresponding species-specific standard and 125I-labeled tracer. There was no crossreactivity with other neuropeptides, gastrointestinal peptides, or hypothalamic extracts of other species. The hypothalamic content in fmol/mg (wet weight) of tissue was 3.6 +/- 0.2 for the human and 11.1 +/- 5.5 for the rat. Age-related changes in hypothalamic GRF content were present in rats, with a gradual increase from 2 to 16 weeks and a correlation between increasing body weight and GRF content. These radioimmunoassays will serve as important tools for understanding the regulation of growth hormone secretion in both human and rat.

The effects of centrally administered neuropeptides on the development of gastric lesions in the rat.

Centrally administered neuropeptides were investigated for their effects on the development of gastric lesions in rats. Thyrotropin releasing hormone (TRH), vasoactive intestinal peptide (VIP) and gonadotropin releasing hormone (LHRH) produced gastric lesions acutely, with TRH demonstrating the most pronounced effect in terms of incidence and severity. Ten-fold higher doses of the same peptides administered intravenously produced none or very few gastric lesions. Moreover, pretreatment with atropine partially inhibited their production. Corticotropin releasing factor (CRF) exhibited only mild ulcerogenic effects, and the gastric lesions induced with this peptide developed more slowly than with TRH, VIP and LHRH. Although ulcerogenic in their own right, none of these four neuropeptides significantly potentiated the potent ulcerogenic effects of cold-restraint stress. Since other neuropeptides, including somatostatin, human pancreatic growth hormone releasing factor (hpGRF), substance P, bombesin, and neurotensin, had no demonstrable effects on gastric mucosa, we can conclude that the lesions were not a general effect of intracisternal administration of neuropeptides. The results suggest that within the central nervous system, there are several neuropeptides that play a significant role in the development of gastric lesions via, at least in part, vagal-dependent mechanisms.

Evidence for a role of endogenous corticotropin-releasing factor in cold, ether, immobilization, and traumatic stress.

The role of corticotropin-releasing factor (CRF) in four model stresses (cold, ether, immobilization, and trauma) was examined in the guinea pig by using passive immunoneutralization with anti-CRF antiserum. Plasma corticotropin levels were measured at various times after exposure to stress, and groups treated with CRF antiserum were compared with those treated with normal rabbit serum. Of the four stresses tested, ether had the most pronounced effect on corticotropin secretion. Treatment with anti-CRF inhibited most of the ether-induced corticotropin secretory response, the difference between the normal serum- and the anti-CRF antiserum-treated groups being significant at 5 and 10 min (P less than 0.01). Corticotropin responses to cold stress in the two groups differed at the 0.05 level of significance at 10 and 20 min. After administration of trauma (leg fracture), a statistically significant difference (P less than 0.01) between the two groups also was evident, albeit only at 20 min. During immobilization, corticotropin levels differed significantly from control only in the normal serum-treated group but not in the anti-CRF-treated group. These findings show that CRF antiserum was effective in reducing corticotropin levels, indicating that CRF has an important role in mediating corticotropin response to stress. The fact that neutralization was incomplete might be due to an inability of the antiserum to sufficiently neutralize the endogenous CRF or, more likely, reflects the contribution of additional mediators, notably catecholamines and vasopressin, of corticotropin release upon stress.

Pituitary adenomas in acromegalic patients: an immunohistochemical and endocrinological study with special reference to prolactin-secreting adenoma.

To establish a functional classification of purification of pituitary adenomas in acromegalic patients, we used immunoperoxidase-staining techniques specific for GH and PRL. Surgical specimens from 55 acromegalic patients were studied. GH was demonstrated in all adenomas and PRL was found in 25 of them (45.5%). Immunohistologically, GH- and PRL-containing adenomas could be divided into 3 types. In type 1 (11 patients), immunoreactive PRL was present in single cells surrounded by immunoreactive GH cells. In type 2 (6 patients), immunoreactive PRL cells formed clusters. In Type 3 (8 patients), immunoreactive PRL and GH cells demonstrated a mosaic pattern, and it was difficult to determine which were in the majority. A double immunostaining method revealed 15 adenomas in which individual cells contained both GH and PRL. Hyperprolactinemia was present in 21 patients, 15 of these had immunoreactive PRL cells (type 1, 4 patients; type 2, 3 patients; type 3, 8 patients). There was no correlation between the size of the adenoma and its type. Endocrinologically, all patients with type 2 and 3 adenomas had an abnormal serum GH response to TRH administration; all type 3 patients had a substantial serum PRL response to TRH administration. Type 3 is considered to be an actively PRL-secreting adenoma, resulting in hyperprolactinemia.

Distribution, biosynthesis, and physiological role of corticotropin-releasing factor in the human: an overview.

Our findings to date indicate that: A peptide resembling oCRF is present in human and mammalian hypothalamus. oCRF is present in human lumbar cerebrospinal fluid. oCRF concentrations do not differ in CSF from normal individuals and from patients with Cushing's syndrome. oCRF appears to be synthesized via a large oligopeptide precursor. An oCRF-like molecule (oCRF-LI) is present in hypothalamic brain tissue. We have also observed more tentative evidence of low levels of oCRF-LI outside of the brain. oCRF is likely to be a central mediator of stress in its multiple forms. We believe that oCRF is clearly of major physiological importance, but that many critical unanswered questions remain. Probably, the most fascinating of these, which we are only beginning to comprehend, concerns the functions of CRF in extrahypothalamic brain as well as the CRF which appears to be present outside the brain.

Anterior pituitary function after transsphenoidal selective adenomectomy in patients with Cushing's disease.

The anterior pituitary function in 25 patients with Cushing's disease was assessed before and after transsphenoidal adenomectomy. Pituitary adenoma was detected and removed in 24 cases, resulting in clinical remission in 22. Postoperative hypoadrenocorticism was observed in all of the cases with remission, necessitating substitution of glucocorticoid. One case had a recurrence after a year in remission. Plasma ACTH and cortisol rapidly decreased after surgery and remained at subnormal levels. However, diurnal rhythmicity of ACTH and cortisol appeared in 5 of 9 cases within 6 months after surgery and exhibited normal suppressibility in response to low dose dexamethasone. The impaired ACTH response to hypoglycemia was restored after surgery. The GH response to hypoglycemia and the TSH response to TRH were improved by correction of hypercorticism and became evident over time. These results suggest that preoperative impairment of anterior pituitary hormone secretion is secondary to hyperadrenocorticism and that ACTH hypersecretion by a primary pituitary adenoma is the primary etiology in Cushing's disease. We conclude that transphenoidal pituitary exploration should be considered as a first choice of treatment of Cushing's disease because of its high clinical remission rate in association with normalization of other endocrine functions.