Morphological and functional alterations of the hypothalamic-pituitary system in brain death with long-term bodily living. Hypothalamic hormones as well as anterior pituitary hormones were detected in the peripheral plasma after the diagnosis of brain death. It is possible that residual hypothalamic tissue was functioning after satisfying the usual criteria of total brain death. To examine this possibility, endocrinological and morphological alterations of the hypothalamic-pituitary system was evaluated in 28 brain dead patients. Intrinsic ADH was depleted in the plasma shortly after the diagnosis of brain death. Anterior pituitary hormones were initially detected in all patients, but gradually disappeared. The direct TRH (thyrotropin releasing hormone) stimulation to the anterior lobe was responded to well. Morphological studies showed a partial necrosis of the anterior lobe and the preservation of the posterior lobe for as long as a week. These data prove that the pituitary is partially preserved after brain death. LH-RH (luteinizing hormone releasing hormone) was detected in the peripheral plasma of all patients and GRF (growth hormone releasing factor) was detected in half of the patients for as long as 15 days, but autopsy revealed the fact that the brain tissue including the hypothalamus became extensively necrotic after the sixth day of brain death. In order to solve this controversy it is proposed that these hormones originate from extracranial tissues such as pancreas. The detection of hypothalamic hormones after the diagnosis of brain death therefore is not contradictory to the concept of total brain death.


Relapse among alcoholics with phobic and panic symptoms. A group of 35 alcoholics who indicated they had symptoms of phobia, panic, or both (the anxiety problem group) were compared to their matched controls who did not indicate having anxiety problems. Comparisons of relapse rates, reasons for relapse, and rates of emotional problems at six months posttreatment were made. Results showed that although relapse rates were similar between the two groups, significantly more anxious subjects reported relapsing to cope with depression and experiencing problems with nervousness, tension, and anger posttreatment. Implications for treatment and the need for further research are discussed.


[Profile of drug addicts admitted to a hospital detoxification unit during a four-and-a-half year period]. It realize a overall and evolutionarily (6 months periods) study concerning the 1,313 drug dependents that were admitted to the Detoxication Unit of the Hospital Psiquiátrico de Madrid, since October 1985 to March 1990. The sociodemographic, drug dependence history, the Physical and toxicological state at admittance, the inpatient evolution, and the familiar and personal psychopathological history variables were evaluated. It bring out that the 45% are dependents to another drug apart from the heroin, the gradual increase in the mean age and in the use of the cocaine, a decrease in the use of the cannabis, anyway the appearance of the buprenorphine addition. There is a gradual decrease, in the period studied, of the antibodies HIV carrier rates. It shows that there is a relationship between this. The poly drug dependents have more legal incidences, psychopathological personal histories (heroin and alcohol and/or benzodiazepines addiction), use more the i.v. route and the HIV seroprevalence are the greatest.


Habitual dietary intake and insulin sensitivity in lean and obese adults. Studies in rodents have shown that short-term increases in dietary fat result in fat cell enlargement and insulin resistance. In humans, although high-fat diets have been associated with obesity, little is known about the specific metabolic effects of these diets. In this study we explored possible associations between habitual dietary composition and insulin sensitivity. Twenty-two lean and 23 obese subjects were characterized by dietary history (food frequency questionnaire), anthropometrics, oral glucose tolerance, and insulin sensitivity (SI, from the minimal model). As shown previously, body mass index was positively correlated with percent of energy intake as fat (r = 0.47, P = 0.001). Increasing fat intake was also associated with diminished SI (r = -0.41, P = 0.01). In contrast, SI was positively correlated with fiber intake (r = 0.43, P = 0.007). Multivariate analysis confirmed the importance of dietary fiber for SI. We conclude that habitually low dietary fiber intake, along with elevated dietary fat, correlates with diminished SI in otherwise healthy lean and obese subjects.


Cranial CT and MR in the Klippel-Trenaunay-Weber syndrome. This report describes the intracranial CT and MR findings in two cases of Klippel-Trenaunay-Weber Syndrome. The findings are 1) markedly enhancing choroid plexuses, 2) severe cerebral atrophy, 3) cerebral calcifications, and 4) angiomatous leptomeningeal enhancement. The findings may resemble those seen in cases of bilateral Sturge-Weber syndrome. The two diseases should be distinguishable by the external stigmata. The authors raise the question of a spectrum of involvement in the angiodysplasias of Klippel-Trenaunay-Weber syndrome and Sturge-Weber syndrome with considerable overlap.


Evaluation of focal epilepsy: a SPECT scanning comparison of 123-I-iomazenil versus HM-PAO. To establish whether regional disturbances of the benzodiazepine receptor distribution in focal epilepsies can be detected by SPECT using 123-I-Iomazenil. Benzodiazepine receptor imaging was carried out in 10 patients interictally. To be eligible for this study the patients had to have a history of focal seizures and no evidence of routine imaging abnormalities as in CT or MR. The patients were selected on the basis of a regional decreased blood flow in HMPAO SPECT that correlated with the site of a stable unifocal EEG abnormality. Benzodiazepine receptor imaging was performed after intravenous administration of approximately 110 MBq 123-I-Iomazenil using SPECT. A regional reduced activity was found on sequential SPECT series after 30 and 90 min post injection in the receptor study. The brain region with a reduced receptor density was concordant to the pathologic finding in HMPAO SPECT in all patients. For the evaluation of patients with focal epilepsies lomazenil SPECT offers several advantages over HMPAO SPECT. Iomazenil binds specifically to the benzodiazepine receptor complex whereas the exact binding sites of HMPAO are still unknown. In contrast to HMPAO, lomazenil can be used for sequential SPECT examinations that may detect dynamic changes of the receptor complex. For the purpose of benzodiazepine receptor imaging, lomazenil is a suitable ligand in patients with focal epilepsies.


A histiocyte-specific marker in the diagnosis of malignant fibrous histiocytoma. Use of monoclonal antibody KP-1 (CD68). KP-1 (CD68) is a recently described monoclonal antibody to a cytoplasmic epitope present on tissue histiocytes and macrophages. To determine the specificity and sensitivity of this marker in the evaluation of cases of malignant fibrous histiocytoma (MFH), this reagent and a panel of commercially antibodies were used to stain formalin-fixed paraffin sections from 25 cases of MFH and 25 other tumors, including a variety of soft-tissue sarcomas. Eighteen of 25 cases of MFH stained for KP-1 (72%), whereas all other tumors were negative, including 12 cases of pleomorphic soft-tissue sarcoma other than MFH. The percentage of tumor cells staining for KP-1 varied. In 11 cases KP-1 was only focally present, but staining was of a high intensity and associated with minimal nonspecific or background staining. Pleomorphic histiocytic cells and spindle cells from storiform tumors were strongly decorated with antibodies to KP-1 in most cases, and antigen also was present on tumor giant cells. Although alpha-1-antitrypsin and alpha-1-chymotrypsin stained a higher percentage of cases of MFH (92%), immunoreactivity for these markers also was noted in other tumors. Because of its specificity as a histiocyte marker, KP-1 is a useful component in a panel of antibodies for the characterization of soft-tissue sarcomas and the diagnosis of MFH.


Large cell carcinoma of the lung with neuroendocrine differentiation. A comparison with large cell "undifferentiated" pulmonary tumors. Twelve large cell carcinomas of the lung showing evidence of neuroendocrine differentiation (LCCND) were compared with 15 other large cell pulmonary tumors that lacked such features (NELCUC). All lesions were composed of partially necrotic, nested, or sheet-like arrays of mitotically active, nucleolated large cells that were at least twice the size of those seen in small cell carcinomas. Examples of LCCND were defined by immunoreactivity for neuron-specific enolase, Leu-7, synaptophysin, and chromogranin-A, and by their content of neurosecretory granules on electron microscopy. NELCUCs were devoid of these immunohistologic and ultrastructural features. There were six women and six men with LCCND, who ranged in age from 38 to 82 years. Of nine individuals in this group with Stage T1NOMO or T2NOMO disease at diagnosis, five (55%) died of their neoplasms within 3 years of diagnosis; three more (33%) have recurrent or persistent tumors and are likely to die as a result. On the other hand, 47% of patients with NELCUC of similar stages are free of disease after a similar follow-up period. LCCND is a distinctive clinicopathologic disease and should not be classified with unspecified large cell anaplastic carcinomas of the lung. Its behavior is potentially aggressive and may justify consideration of a specialized treatment protocol. Because electron microscopic evaluation of immunohistologic features must be done to recognize LCCND, it is probably underdiagnosed.


Russell bodies consist of heterogenous glycoproteins in B-cell lymphoma cells. The phenotypic expression of Russell bodies (RB) in the tumor cells of two patients with different types of B-cell lymphoma and of one patient with plasma cell myeloma were examined. In both B-cell lymphomas, the RBs reacted consistently with anti-Leu 8 and anti-immunoglobulin M. The RBs in one case also reacted with other monoclonal antibodies, including anti-CD5, CD19, CD22, and CD25. The membranes of most of the tumor cells containing RBs did not stain. In the myeloma, the RBs reacted only with anti-immunoglobulin, and the myeloma cells expressed no surface antigens associated with B lymphocytes. This finding suggests that RBs do not form in cells that can transport glycoproteins to the cell membrane, but rather occur as a result of defective transport or process of certain glycoproteins by plasmacytoid cells.


Evaluation of sexually abused and nonabused young girls for intravaginal human papillomavirus infection. The objective of this study was to compare the prevalence of intravaginal human papillomavirus-associated disease in two groups of girls to develop information regarding the means of transmission of anal-genital human papillomavirus disease. A pair of parallel studies of prevalence of human papillomavirus infections in two populations of prospectively enrolled girls. Index patients consisted of 15 consecutive girls aged 11 years or younger who were confirmed to have been sexually abused, had signs or symptoms of vaginal disease, and required generalized anesthesia for evaluation. Selection of nonabused control patients was based on negative findings from screening evaluations and physical examinations. Prevalences of cervical-vaginal human papillomavirus infections in the two populations were compared. Vaginal wash samples from index and control patients were assayed for human papillomavirus 1, 2, 4, 6, 11, and 16 by reverse-blot and Southern transfer hybridization methods. Papanicolaou smears were examined from index patients. Vaginal wash samples from five (33%) of 15 index patients were positive for human papillomavirus 6, 11, or 16, compared with none of 17 controls. The presence or absence of external anal-genital warts was not correlated with results from the assay of intravaginal samples. Blinded readings of vaginal exfoliative cytologic findings of the index patients showed koilocytosis, atypia, or inflammatory reactions in four of five human papillomavirus-positive girls, and normal cytologic findings in one human papillomavirus-positive girl. These findings support other studies that indicate that sexual contact is a major route in the transmission of anal-genital human papillomavirus-related disease in children. Evaluation of intravaginal specimens was required to identify human papillomavirus-infected girls since the results of the wash samples were not correlated with the presence or absence of external anal-genital warts.


Intraductal mucin-producing tumors of the pancreas. Four cases of intraductal mucin-producing tumors of the pancreas are presented, with a literature review. These tumors have been reported with increasing frequency, especially in Japan, since it was advocated that they represented a new clinical entity in 1982. Despite the recent controversy, intraductal mucin-producing pancreatic tumor has been recognized as a distinct entity, i.e., mucinous ductal ectasia. In the present paper, clinical features of these tumors and their relation to our own classification of mucin-producing pancreatic tumors are discussed.


Small cell carcinoma of the esophagus in a patient with longstanding primary achalasia. Achalasia is believed to be a predisposing factor for the development of esophageal cancer. Small cell carcinoma of the esophagus is a rare neoplasm, with fewer than 150 cases having been reported in the world literature, and it has been described only once previously in a patient with longstanding achalasia. We describe a case of an 85-yr-old woman with long-term primary achalasia who developed primary small cell carcinoma of the esophagus. We hypothesize that this patient's recurrent, worsening dysphagia is related to a paraneoplastic phenomenon. We discuss this association and review the literature.


Case report: severe symptomatic hyponatremia associated with lisinopril therapy. A 63-year-old white female who was being treated for hypertension with lisinopril presented with seizures, altered mental status, and a serum sodium of 101 mEq/L. Serum sodium prior to initiation of lisinopril therapy was 137 mEq/L. The hyponatremia was corrected and did not recur after lisinopril was stopped. The hyponatremia may have been a result of polydipsia and inappropriate antidiuresis secondary to ACE-inhibitor therapy.


Lack of reactivity of uterine arteries from patients with obstetric hemorrhage. Obstetric hemorrhage may occur throughout pregnancy and the puerperium. The purpose of this study was to investigate the reactivity of isolated, suffused uterine arteries from obstetric patients with uncontrollable uterine bleeding and to compare those blood vessels with uterine arteries from patients undergoing cesarean hysterectomy for other medical reasons (control patients). The uterine arteries from the control patients (n = 9) responded with maximal or near-maximal constriction to norepinephrine (30 mumol/L, 3.6 +/- 1 gm), potassium chloride (75 mmol/L, 10.2 +/- 3 gm), prostaglandin F2 alpha (30 mumol/L, 1.8 +/- 1 gm), and arginine vasopressin (1 mumol/L, 18.8 +/- 2.6 gm). In uterine arteries from five patients with uncontrollable bleeding, the constrictor responses to the same drugs were markedly depressed: norepinephrine (30 mumol/L, 0.5 +/- 0.2 gm), potassium chloride (75 mmol/L, 1.9 +/- 0.8 gm); prostaglandin F2 alpha (30 mumol/L, 0 gm), and arginine vasopressin (1 mumol/L, 0.2 +/- 0.05 gm). Uterine arteries from two patients exhibited no constrictor responses to norepinephrine (30 mumol/L), potassium chloride (75 mmol/L), prostaglandin F2 alpha (30 mumol/L), or arginine vasopressin (1 mumol/L). The impaired responses to the vasoconstrictor drugs were not reversed by indomethacin (1 mumol/L), which is an inhibitor of prostaglandin synthetase; methylene blue (10 mumol/L), which is a blocker of endothelium-derived relaxing factor activation of guanylate cyclase; or propranolol (1 mumol/L), a beta-adrenergic receptor antagonist. The levels of adenosine 3':5'-cyclic monophosphate were not elevated in the uterine arteries from the patients with obstetric hemorrhage. The impaired reactivity to the multiple vasoconstrictors implies that a mechanism involved in constriction common to all of the constrictors is depressed or blocked. Furthermore, the depression or lack of reactivity of these isolated uterine arteries is not mediated by vasodilatory prostaglandins, endothelium-derived relaxing factor, beta-adrenergic receptors, or elevated levels of adenosine 3':5'-cyclic monophosphate. The results suggest that obstetric hemorrhage involves, in part, a lack of constrictor reactivity of the uterine vasculature.


Origin of cervical collagenase during parturition. Cervical biopsy specimens were obtained under standard conditions from the posterior lip of the uterine cervix in 105 patients. A significant increase of collagenase activity was observed during parturition as determined with an assay with iodine 125-labeled native triple-helical collagen type I as the substrate. The collagenase was not likely to originate from cervical fibroblasts because in situ hybridization failed to detect synthesis of the specific procollagenase messenger ribonucleic acid. However, migration of polymorphonuclear leukocytes into the cervical stroma occurred on onset of labor, and an antibody specific for human leukocyte collagenase that did not cross react with fibroblast collagenase revealed the presence of the enzyme in the granules of polymorphonuclear leukocytes and subsequently in the extracellular matrix of the cervix. Therefore it is likely that the cells critically involved in collagen degradation during cervical dilatation are not resident fibroblasts but rather polymorphonuclear leukocytes emigrating from blood vessels.


Influence of labor and oxytocin on in vitro leukotriene release by human fetal membranes and uterine decidua at term gestation. The purpose of this study was to determine whether labor and oxytocin influence in vitro leukotriene release by human fetal membranes and uterine decidua at term gestation. All women (N = 21) were studied in an academic clinical research environment. They were delivered either vaginally or before labor by cesarean section. Tissues were incubated with calcium ionophore A23187 alone or with calcium ionophore A23187 after preincubation with either oxytocin, BW755c, or cycloheximide. Leukotrienes in culture medium were determined by high-performance liquid chromatography. Statistical analysis was performed with analysis of variance. Tissues obtained after vaginal delivery released significantly higher (p less than 0.05) quantities of leukotrienes LTB4, LTD4, and delta-6-trans-LTB4 than did tissues obtained during cesarean section. Leukotriene release was stimulated by oxytocin and was inhibited by both BW755c and cycloheximide. These results suggest an activation of the arachidonate lipoxygenase pathway in these tissues during labor. Oxytocin could play a regulatory role in this process.


Semisynthesis and purification of homogeneous plasmenylcholine molecular species. Methods for the efficient use of limiting amounts of fatty acid probes in the synthesis of individual molecular species of plasmenylcholine have been developed. Plasmenylcholine molecular species were synthesized through acylation of homogeneous 1-O-(Z)-hexadec-1'-enyl-sn-glycero-3-phosphocholine utilizing fatty acid anhydrides generated in situ from combined pools of reactant and recycled fatty acids by repeated addition of small amounts (10 mol%) of N,N'-dicyclohexylcarbodiimide. The efficient generation of reactive anhydrides was accomplished through minimizing irreversible formation of N-acyl urea adducts by maintaining a persistent molar excess of fatty acid (with respect to carbodiimide) during the entire reaction time course. The synthesis of multiple different sn-2 labeled plasmenylcholine probes for utilization in fluorescence, ESR, or 2H NMR spectroscopy as well as isotopically labeled plasmenylcholines for metabolic studies has been achieved in good yield (40-50% of theoretical yield based on fatty acid) by these methods. Rapid and effective purification methods utilizing high-performance liquid chromatography were developed for both large- and small-scale purifications of individual reaction mixtures which collectively resulted in the isolation of homogeneous plasmenylcholine molecular species in high yield from limiting amounts of fatty acid probes.


Selective determination of arginine-containing and tyrosine-containing peptides using capillary electrophoresis and laser-induced fluorescence detection. The use of two different amino acid-selective fluorogenic reagents for the derivatization of peptides is investigated. One such scheme utilizes a selective reaction of benzoin with the guanidine moiety to derivatize arginine residues occurring in a peptide. The second scheme involves the formylation of tyrosine, followed by reaction with 4-methoxy-1,2-phenylenediamine. The use of capillary electrophoresis and laser-induced fluorescence detection allows enhanced efficiencies and sensitivities to be obtained for the separations of either arginine- or tyrosine-containing peptides. A helium-cadmium laser (325 nm) is ideally suited for the laser-based detection system due to a close match of the excitation maxima of derivatized peptides from both reactions. A detection limit of 270 amol is achieved for model arginine-containing peptides, while the detection limit for model tyrosine-containing peptides is measured at 390 amol. Both derivatization reactions are found to be useful for high-sensitivity peptide mapping applications in which only the peptides containing the derivatized amino acids are detected.


Cytochrome c aided resolution of Lupinus albus isoperoxidases in a cathodal polyacrylamide gel electrophoresis system. In a cathodal polyacrylamide gel electrophoresis system, three distinct groups of isoperoxidases from Lupinus albus were found to achieve retention factors (rf) dependent on the quantity of sample applied onto the gel. The possibility of extract-derived substances weakly associating with peroxidase samples was investigated. Association of the putative agents survived dialysis against electrophoresis buffer with and without 2 M CaCl2 and freeze-thaw treatments. The addition of polyvinylpolypyrrolidone and polyethylene glycol to the homogenization buffer also proved ineffective in eliminating the variation in isoperoxidase rf although differences in the zymogram profiles of these samples were evident. The addition of spermine and cytochrome c to samples was found to increase the rf of some peroxidase bands. Electrophoresis of samples with cytochrome c resulted in the resolution of peroxidase groups to distinct bands at rf independent of the quantity of peroxidase applied. Control experiments indicate that this treatment did not introduce any detectable artifacts.


Mass spectrometry of high-mannose oligosaccharides after trifluoroacetolysis and periodate oxidation. High-mannose oligosaccharides were treated with a mixture of trifluoroacetic acid and trifluoroacetic anhydride under conditions where reducing terminal N-acetylglucosamine residues are specifically degraded. The resulting mannose-containing products were, after further chemical modifications, analyzed by mass spectrometry in fast atom bombardment and electron ionization modes. Binding positions between monosaccharide residues were deduced from mass spectra of peracetylated compounds, which, prior to the derivatization, had been subjected to periodate oxidation and borodeuteride reduction.


Pharmacokinetics and pharmacodynamics of pipecuronium bromide (Arduan) in elderly surgical patients. The neuromuscular response to pipecuronium bromide (Arduan), 70 micrograms/kg, was studied in 20 elderly (greater than 70 yr) and 10 younger patients (less than 60 yr) during nitrous oxide, fentanyl, and droperidol anesthesia. The adductor pollicis response to single 0.2-ms supramaximal pulses was recorded. Although all younger patients were completely paralyzed, 2 of 20 elderly patients did not attain 90% paralysis. Onset time in the elderly was prolonged (6.9 +/- 2.6 vs 4.3 +/- 1.5 min, P less than 0.02). Spontaneous recovery was similar in both groups, with 75% recovery occurring at 133 +/- 52 min in the elderly and 146 +/- 46 min in the younger patients. The pharmacokinetic variables were similar for the two groups, and pharmacodynamic analysis revealed a similar sensitivity at the neuromuscular junction. The pharmacologic actions of pipecuronium in otherwise healthy patients do not differ between young and elderly adults.


Evaluation of the antihypertensive effect of lisinopril compared with nifedipine in patients with mild to severe essential hypertension. For several reasons, increasing numbers of patients with hypertension are treated with angiotensin-converting enzyme inhibitors and calcium channel blockers. In a twenty-four week, double-blind, randomized, parallel study, the antihypertensive effect of lisinopril (20 to 80 mg qd) and nifedipine (20 to 80 mg bid) were compared in 21 patients. Fourteen patients received lisinopril (mean dose 35 mg), and 7 patients received nifedipine (mean dose 54 mg). By the end of week 12, 8 patients had responded (supine diastolic pressure less than or equal to 90 mg) to lisinopril and 5 to nifedipine. At the end of the study supine systolic/diastolic blood pressure was reduced from 172/104 to 149/92 mmHg with lisinopril and from 171/102 to 158/94 mmHg with nifedipine. No significant difference between the two treatments was detected. Three patients were reported to have at least one clinical adverse experience during the active treatment period, 1 in the lisinopril group and 2 in the nifedipine group. No serious clinical adverse experiences were recorded. In conclusion, lisinopril and nifedipine are both effective in reducing blood pressure in patients with mild to severe hypertension. Lisinopril qd and nifedipine slow release bid produce similar decreases in blood pressure after twelve weeks of therapy and the safety profiles of the two drugs are similar.


Metabolic response of horses to a high soluble carbohydrate diet: effects of low-intensity submaximal exercise and sodium bicarbonate supplementation. Four mares fed a low fiber, high soluble carbohydrate diet were used in a crossover design to evaluate the effects of dietary sodium bicarbonate (NaHCO3) supplementation during daily low-intensity submaximal working conditions. Mares were fed the diet at 1.7 times the maintenance energy requirement for mature horses at work. The horses tolerated the diet well and had no clinical abnormalities. Resting venous blood bicarbonate (HCO3), standard HCO3, and base excess (BE) concentrations significantly (P less than 0.05) increased with NaHCO3 supplementation, but no significant changes in resting venous blood pH or carbon dioxide tension (PCO2) were recorded. Venous blood HCO3, standard HCO3, BE, hemoglobin, and heart rate were significantly (P less than 0.05) increased and plasma lactate concentration was significantly (P less than 0.05) decreased in the control horses and in the horses given the NaHCO3 supplement during low-intensity submaximal exercise. There were no significant changes in venous blood pH, PCO2, or plasma protein concentration with exercise. Venous blood HCO3, standard HCO3, and BE concentrations were significantly (P less than 0.05) greater during submaximal exercise in horses given the NaHCO3 supplement. There were no significant differences in plasma lactate or total protein concentrations, blood pH, PCO2, or hemoglobin concentration between the 2 groups during exercise.


Inoculation of pigs with Streptococcus suis type 2 alone or in combination with pseudorabies virus. Pigs (9 [+/- 1] weeks old) were inoculated with Streptococcus suis type 2, pseudorabies virus (PRV), or both. For each pig of groups A, B, and C the inoculum of S suis was 10(9) colony-forming units. For each pig of groups A, B, and D the inoculum of PRV was 5 x 10(3) TCID50 of either PRV strain 4892 (group A, n = 9) or PRV isolate B (group B, n = 9). The PRV strain 4892 is a highly virulent strain; isolate B causes mild clinical signs of infection in inoculated pigs. Group-C pigs (n = 9) were given S suis alone, and group-D pigs (n = 3) were inoculated only with PRV isolate B. Clinical signs of infection and development of lesions were readily seen in pigs of groups A, B, and C. Duration and severity of clinical signs of disease and lesions were reduced in pigs of group C, compared with those of the other 2 groups. Lesions, such as polyarthritis and fibrinous pericarditis, were more abundant and acute in the groups of pigs given mixed challenge exposure, compared with pigs inoculated exclusively with S suis type 2 (group C). The group of pigs inoculated with PRV isolate B alone did not manifest clinical signs of disease or lesions. Average daily gain for group-C pigs was higher, compared with that of other groups; the difference was statistically significant at P less than 0.02 and P less than 0.05 for groups B and D, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)


Effects of synthetic ovine corticotropin-releasing hormone on plasma concentrations of immunoreactive adrenocorticotropin, alpha-melanocyte-stimulating hormone, and cortisol in dogs with naturally acquired adrenocortical insufficiency. We evaluated the effect of ovine corticotropin-releasing hormone (CRH) on plasma immunoreactive (IR) concentrations of ACTH, alpha-melanocyte-stimulating hormone, and cortisol in 8 dogs with naturally acquired adrenocortical insufficiency. Of the 7 dogs with primary adrenal insufficiency, 6 had markedly high basal plasma IR-ACTH concentrations and exaggerated ACTH responses to CRH administration, whereas 1 dog that was receiving replacement doses of prednisone at the time of testing had normal basal IR-ACTH concentrations and a nearly normal response to CRH. In contrast, the 1 dog with secondary adrenocortical insufficiency had undetectable basal plasma IR-ACTH concentrations, which failed to increase after administration of CRH. Basal plasma alpha-melanocyte-stimulating hormone concentrations in the dogs with adrenal insufficiency were within normal range and were unaffected by CRH administration. In all 8 dogs with adrenal insufficiency, plasma cortisol concentrations were low and did not increase after administration of CRH. Therefore, stimulation with CRH produced 2 patterns of plasma IR-ACTH response when administered to dogs with naturally acquired adrenal insufficiency. Dogs with primary adrenal insufficiency had high basal plasma IR-ACTH concentrations and exaggerated responses to CRH, whereas the dog with secondary adrenal insufficiency had undetectable basal plasma concentrations of IR-ACTH that did not increase after stimulation with CRH.


The effect of an inhaled neutral endopeptidase inhibitor, thiorphan, on airway responses to neurokinin A in normal humans in vivo. Neuropeptides such as neurokinin A (NKA) have been proposed as important mediators of bronchoconstriction and airway hyperresponsiveness in asthma. Inhaled NKA causes bronchoconstriction in patients with asthma, but not in normal subjects. This is possibly due to the activity of an endogenous neuropeptide-degrading enzyme: neutral endopeptidase (NEP). We investigated whether a NEP-inhibitor, thiorphan, reveals bronchoconstriction to NKA or NKA-induced changes in airway responsiveness to methacholine in normal humans in vivo. Eight normal male subjects participated in a double-blind crossover study, using thiorphan as pretreatment to NKA challenge. Dose-response curves to inhaled NKA (8 to 1,000 micrograms/ml, 0.5 ml/dose) were recorded on 2 randomized days 1 wk apart, and methacholine tests were performed 48 h before and 24 h after the NKA challenge. Ten minutes prior to NKA challenge the subjects inhaled either thiorphan (2.5 mg/ml, 0.5 ml) or placebo. To detect a possible nonspecific effect of thiorphan, we investigated the effect of the same pretreatment with thiorphan or placebo on the dose-response curve to methacholine in a separate set of experiments. The response was measured by the flow from standardized partial expiratory flow-volume curves (V40p), expressed in percent fall from baseline. NKA log dose-response curves were analyzed using the area under the curve (AUC) and the response to the highest dose of 1,000 micrograms/ml (V40p,1000). The methacholine dose-response curves were characterized by their position (PC40V40p) and the maximal-response plateau (MV40p). Baseline V40p was not affected by either pretreatment (p greater than 0.15).(ABSTRACT TRUNCATED AT 250 WORDS)


Cellular mechanisms that control pulmonary vascular tone during hypoxia and normoxia. Possible role of Ca2+ATPases. We investigated cellular mechanisms that may be involved in controlling cytosol calcium and pulmonary artery pressure during hypoxia and normoxia in isolated blood-perfused ferret lungs. Alveolar hypoxia in ferret lungs causes an active increase in pulmonary vascular resistance. Hypoxic pulmonary vasoconstriction directly correlates with extracellular calcium ([Ca2+]o), and the absence of [Ca2+]o in the perfusate markedly attenuates the hypoxemia-induced pulmonary vasoconstriction. Alveolar hypoxia does not potentiate the production of thromboxane B2 (TxB2) or 6-keto-PGF1 alpha. Vanadate, a widely used inhibitor of Ca2+ATPases, increases pulmonary arterial pressure (Ppa) in the presence or absence of [Ca2+]o and without affecting the production of TxB2 or 6-keto-PGF1 alpha. Vanadate and ouabain, an inhibitor of Na+/K+ATPase, produce synergistic increases in Ppa. Amiloride, an inhibitor of Na+/Ca2+ exchange, reverses the increase in Ppa caused by ouabain, but not the increase caused by vanadate. The additional effect produced by ouabain on Ppa after near maximal vanadate effect and the ability of amiloride to reverse the pulmonary vasoconstriction caused by ouabain, but not vanadate, suggests that vanadate does not inhibit Na+/K+ATPase in ferret lungs. In addition, cyclic GMP (cGMP), which has been reported to increase the activity of Ca2+ATPases in vascular smooth muscle, was able to reverse and prevent the effect of vanadate on Ppa, but not the effect of ouabain. Inhibition of Ca2+ATPases with vanadate in ferret lungs increases pulmonary vascular resistance during both normoxia and hypoxia. The Ca2+ entry mediated by alveolar hypoxia appears to overpower the ability of Ca2+ATPases and other membrane Ca2+ transport proteins to translocate [Ca2+]i.(ABSTRACT TRUNCATED AT 250 WORDS)


Recombinant-human superoxide dismutase attenuates endotoxin-induced lung injury in awake sheep. Oxygen radicals have been implicated in the pathogenesis of acute lung injury associated with clinical and experimental sepsis. With the use of endotoxin infusion as an in vivo model of sepsis we studied the effect of recombinant-human superoxide dismutase (r-hSOD; 4,200 U/mg), an enzyme that catalyzes the dismutation of superoxide anion, on both the physiologic and biochemical lung changes in awake sheep. Sheep (n = 11) were prepared for chronic measurement of pulmonary hemodynamics and lung fluid balance. Paired experiments were performed in seven of the animals in which they received either endotoxin (1 microgram/kg) alone or in combination with r-hSOD in random order. An additional four sheep received r-hSOD without the lipopolysaccharide. Intravenous infusion of r-hSOD (a loading dose of 12,600 U/kg followed by a maintenance dose of 14,700 U/kg/h for 7 h) resulted in substantial SOD activity, measured by electron spin resonance spectrometry, both in plasma and in lung lymph, and attenuated the expected changes in pulmonary arterial pressure and lung lymph flow after administration of endotoxin. When administered without endotoxin, r-hSOD produced no perceptible change in pulmonary hemodynamics and lung fluid balance. These data suggest that superoxide anion plays an important role in endotoxin-induced lung injury in sheep.


Effects of naproxen on experimental rhinovirus colds. A randomized, double-blind, controlled trial. To determine whether naproxen, a propionic acid inhibitor of cyclooxygenase, alters the course of experimental rhinovirus colds. A randomized, double-blind, controlled trial. Rhinovirus challenge model in volunteers cloistered in individual hotel rooms. Eighty-seven healthy young adults with serum neutralizing antibody titers of less than or equal to 1:2 to the challenge virus; 79 were evaluable. Thirty-nine participants received naproxen (loading dose, 400 mg or 500 mg followed by 200 mg or 500 mg three times daily for 5 days). Forty participants received placebo. Treatment was started 6 hours after viral challenge. Daily measurement of viral titers, symptoms, nasal mucus production, and nasal tissue use; incidence of infection and illness; and measurement of homotypic serum neutralizing antibody responses. Viral titers and serum homotypic antibody responses were similar in the naproxen and placebo groups. Significant reductions in headache, malaise, myalgia, and cough occurred in the naproxen group. A 29% reduction was noted in the total (5-day) symptom score in the naproxen group (95% CI, 16% to 42%). Naproxen treatment did not alter virus shedding or serum neutralizing antibody responses in participants with experimental rhinovirus colds, but it had a beneficial effect on the symptoms of headache, malaise, myalgia, and cough. Prostaglandins may be among the inflammatory mediators that play a role in the pathogenesis of rhinovirus colds.


Shave excision and dermabrasion for facial angiofibroma in tuberous sclerosis. Tuberous sclerosis is an inheritable disease of varied manifestations. Hallmarks of the disease have historically been identified as infantile seizures, severe mental retardation, and facial growths. The facial lesions were formerly termed adenoma sebaceum, but are now known to be angiofibroma. We present a patient who was referred for management of large facial lesions complicated by intermittent hemorrhage. A combination of shave excision and dermabrasion led to a symptomatic and cosmetic improvement.


Idiopathic recurring stupor: a case with possible involvement of the gamma-aminobutyric acid (GABA)ergic system. A patient had recurrent spontaneous episodes of stupor or coma in the absence of toxic, metabolic, or structural brain damage. Ictal electroencephalography showed fast 14 Hz background activity; sleep studies excluded narcolepsy. Flumazenil (Anexate), a benzodiazepine antagonist, promptly resolved the episodes and normalized the electroencephalogram. Radioreceptor binding studies showed the presence of a ligand to the central benzodiazepine receptor in plasma and cerebrospinal fluid during the episodes, suggesting a gamma-aminobutyric acid (GABA)ergic system involvement in the origin of the attacks.


Randomized trial of neoadjuvant therapy for lung cancer: interim analysis. The role of neoadjuvant chemotherapy in stage IIIa non-small cell lung cancer remains undefined. Since 1987, 27 patients with non-small cell lung cancer, all with histologically confirmed metastases to the ipsilateral mediastinal lymph nodes, have been enrolled in an ongoing prospective, randomized trial at our institution. Thirteen patients have been randomized to preoperative etoposide-platinum (EP) chemotherapy-surgery-postoperative EP, and 14 other patients have been randomized to surgery-postoperative mediastinal irradiation (SRT). Both groups are similar in sex, age, weight loss, tumor location, preoperative pulmonary function, physiologic grade, and tumor histology. Eight of the 13 EP patients have responded as evidenced by a 50% or greater radiographic tumor shrinkage after two cycles. Complete tumor and nodal resection rates were similar: 11/13 EP patients versus 12/14 SRT patients. There was no operative mortality for the 27 patients. Median potential follow-up is 29.9 months for the EP group and 34.9 months for the SRT group. Preliminary results suggest a trend toward increased survival time for the EP group (median, 28.7 months) versus the SRT group (median, 15.6 months) (p2 = 0.095). Eleven of 12 resected SRT patients have had recurrence versus 8 of 11 resected EP patients. Time to recurrence reveals no significant differences between the two groups but a trend toward increased disease-free interval in the EP group (12.7 months versus 5.8 months, EP versus SRT). This interim analysis demonstrates the feasibility of such a trial; however, despite the trends, definitive conclusions await further accrual and study maturation.


Papillomavirus found in anorectal squamous carcinoma, not in colon adenocarcinoma. We used polymerase chain reaction DNA amplification methods for the detection and typing of genital human papillomaviruses in paraffin-embedded tissue sections of five patients with anorectal squamous cell carcinoma and 22 patients with colonic adenocarcinoma. The cases were further tested by in situ hybridization with biotin-labeled probes specific for human papillomavirus types 6/11, 16/18, and 31/33/35. By polymerase chain reaction, human papillomavirus DNA was demonstrated in all of the cases of anorectal squamous cell carcinoma and in none of the cases of colonic adenocarcinoma for which analyzable DNA was available. Tumor cell nuclei stained for human papillomavirus DNA by in situ hybridization in four of the five cases of squamous cell carcinoma and in none of the cases of colonic adenocarcinoma. We conclude that human papillomavirus types usually associated with malignant transformation are uniformly present in anorectal squamous cell carcinoma but are absent from adenocarcinoma of the colon.


[Intrauterine infections: frequency and diagnosis]. Intrauterine infections occupy a considerable place in human pathology--a conclusion drawn on the basis of original and literature data. Herpetic, cytomegalic, mycoplasma and chlamydial infections are the most important among them. Intrauterine infections may be the cause of some diseases considered as somatic ones. The possibility of the pathology diagnosis of these disease not only on the basis of laboratory data but on the basis of characteristic structural changes of cells and tissues is shown.


Subacute idiopathic demyelinating polyradiculoneuropathy. Seven cases of subacute idiopathic demyelinating polyradiculoneuropathy had a monophasic illness characterized by progressive weakness of all four limbs that evolved during 4 to 8 weeks. Neurophysiological investigations implied demyelination in all seven cases. In two patients, sural nerve biopsy specimens that were taken showed macrophage-associated demyelination. All patients made substantial or complete recoveries with oral prednisolone (four cases) or without treatment (three cases). None of the patients required ventilation or had autonomic complications. These cases provide a link between the acute idiopathic demyelinating form of Guillain-Barré syndrome and chronic idiopathic demyelinating polyradiculoneuropathy.


The effect of delmopinol on glucosyltransferase adsorbed on to saliva-coated hydroxyapatite. The aim was to explore the effects of delmopinol, a substituted amino-alcohol compound recently reported as a potential antiplaque agent, on GTF activity in solution and when adsorbed on to sHA. Delmopinol was without a significant effect on GTF activity in solution. In contrast, a reduction in the bound glucans synthesized by the adsorbed GTF was found in the presence of delmopinol. Delmopinol did not displace the adsorbed GTF from the sHA, nor was there significant desorption of glucans from sHA. The total glucan synthesis (bound and unbound) was reduced in the presence of delmopinol. Inhibition of GTF was not reversed by sucrose. Inhibition of GTF activity by delmopinol apparently results from drug-enzyme interaction on the surface of sHA beads. These observations provide further support for the important differences in the properties of adsorbed GTF and GTF in solution, illustrating that GTF-drug interaction differs between enzyme adsorbed to surfaces and enzyme in solution.


Relationship between incorporation of choline into phosphatidylcholine and morphine binding sites. Calcium (25 mM)-stimulated incorporation of choline into phosphatidylcholine (PC) and 3H-morphine (3H-M) binding was studied using cerebral crude synaptosomal fractions from acute morphine-treated (naive), morphine-dependent and morphine-withdrawn mice. Mice became physically dependent on morphine after ingesting morphine-mixed food. Morphine dependence inhibited the incorporation of 14C-choline into PC. The injection of naloxone (1 mg/kg) to morphine-dependent mice also inhibited the incorporation of 14C-choline into PC. The injection of morphine (10 mg/kg) to morphine-withdrawn mice stimulated the incorporation of 14C-choline into PC, but had no effect, either on acute morphine-treated mice or on morphine-dependent mice. Naloxone (1 mg/kg), morphine (10 mg/kg), morphine dependence and opiate withdrawal (20 hrs after the last exposure to morphine in food) all caused about a 25% decrease in 3H-M (10(-7) M) binding to mouse brain synaptosomes. The injection of 10 mg/kg of morphine 45 min before death caused an additional 20% decrease in 3H-morphine binding in the dependent and withdrawn mice, but not in the naive mice. Stereospecific binding (SSB) of 3H-M was evaluated with respect to: 1) the effect of naloxone treatment in vivo on 3H-M binding in vitro, 2) the effect of morphine treatment in vivo on 3H-M binding in vitro; and 3) the differences between 3H-M bound (in vitro) and both naloxone and morphine treatment (in vivo). SSB activity was high in morphine-withdrawn mice. These differences may be caused by changes in the opiate receptors related to tolerance and withdrawal, and may entail between PC and membrane receptors.


[The occurrence of virus enteritis (duck plague) in wild birds]. 296 samples from wild birds of 15 species were incorporated into long term exploration of duck plague on ducks in farms. By using virological and serological standard methods 9 virus carriers and 20 serum samples showing positive antibody titers could be detected. The epidemiology as well as the relation of the incidence of duck plague in wild birds and farm poultry is discussed.


Stabilization of the peripheral-type benzodiazepine acceptor by specific phospholipids. The peripheral-type benzodiazepine acceptor from rat adrenal gland was solubilized with Triton X-100. The soluble acceptor exhibited isoquinoline carboxamide and benzodiazepine binding activity only when supplemented with lipid. Phosphatidylserine and phosphatidylinositol were especially efficacious. Such selectivity may reflect a specific structural requirement for acceptor activity. The requirement for lipid for acceptor activity in fractions subsequent to solubilization was demonstrated. This stabilization of acceptor activity by the phospholipids promises to facilitate purification of the functional acceptor.


Ca2+/calmodulin-dependent protein kinase II from hen brain. Purification and characterization. Ca2+/calmodulin-dependent protein kinase II (CaM-kinase II) has been purified from hen whole brain. The enzyme was purified 3000-fold using phosphocellulose and calmodulin-Agarose column chromatography. The specific activity was 200 nmol/min/mg protein. Microtubule associated protein-2 (MAP-2) was used as a substrate to assess the activity of the enzyme during purification and for its characterization. CaM-kinase II consisted of alpha and beta/beta' subunits of molecular weights 46,000 and 55,000/52,000, respectively. The ratio of alpha to beta/beta' subunits was 3:1 in the enzyme purified from the whole brain. The enzyme exhibited broad substrate specificity and phosphorylated myelin basic protein, MAP-2, histone II, histone VIII, casein, tubulin, myosin light chains, glycogen synthase, and phosvitin in decreasing order. Phosphorylase b was phosphorylated at a negligible rate. Autophosphorylation of CaM-kinase II for 10 min in the presence of calcium and calmodulin decreased its total activity to 33%, and calcium/calmodulin-independent activity reached 30% after 1 min and then dropped to 14% after 10 min of autophosphorylation. The Km value of ATP was 19 +/- 1.3 microM, and the K0.5 values of calcium and calmodulin were 4.4 +/- 0.5 and 3.0 +/- 0.5 microM, respectively. The latter were determined using myelin basic protein as the substrate. CaM-kinase II exhibited great differences in the calmodulin requirement for phosphorylation of MAP-2, histone II and myelin basic protein. MAP-2 required the least amount of calmodulin for its phosphorylation. Autophosphorylation of CaM-kinase II resulted in decreased mobility of the alpha-subunit but apparently not of the beta/beta' subunits in sodium dodecyl/sulfate-polyacrylamide gel. Antiserum was raised against the CaM-kinase II alpha subunit and used for testing cross-reactivity of hen brain enzyme with that of other species. The antiserum which reacted with both alpha and beta subunits of hen brain CaM-kinase II cross-reacted with only the alpha subunit of rat, mouse, rabbit, cat, dog, pig and human brain samples. The purified hen brain CaM-kinase II is a multifunctional enzyme and resembled rat brain CaM-kinase II in several properties. Immunocross-reactivity suggested that there was similarity in the alpha but not the beta/beta' subunits of the hen brain enzyme and the brain enzyme of other species.


Effect of ethanol on the Na(+)- and the Na+,K(+)-ATPase activities of basolateral plasma membranes of kidney proximal tubular cells. The Na(+)- and the Na+,K(+)-ATPase activities of basolateral plasma membranes from rat kidney proximal tubular cells were affected differentially by ethanol. Moreover, at concentrations of ethanol that can be reached in vivo in the blood plasma (50 mM) there was a significant effect on the Na(+)-ATPase activity and practically no effect on the Na+,K(+)-ATPase activity.


Cytomegalovirus infection induces vascular injury in the rat. The role of cytomegalovirus (CMV) in the early development of atherosclerosis was studied in a rat model. Arterial samples derived from virus-infected normo- and hypercholesterolaemic animals were investigated by light microscopy at 1, 4, 8 and 16 weeks post infection. Early atherogenic lesions comparable to those seen in non-infected hypercholesterolaemic rats were found in CMV-infected normocholesterolaemic and hypercholesterolaemic animals, starting at 1 week post infection. The changes consisted of minimal endothelial cell damage, as shown by the en face technique, and a more than 10-fold increase in the number of leukocytes adhering to the aortic intima. The increased adhesion of leukocytes was observed in infected normocholesterolaemic rats but only in the non-infected rats which were hypercholesterolaemic. The infection of hypercholesterolaemic rats did not enhance this effect although it resulted in increased migration of the leukocytes into the subendothelial space. CMV infection of normocholesterolaemic rats induced lipid accumulation in the endothelium. In these animals approximately 1% of the endothelial cells contained lipid at 1 week post infection. In the non-infected hypercholesterol-fed animals 10% of the cells contained lipid. CMV infection in these rats induced an extra increase of the lipid-containing endothelial area. The changes in the CMV infected animals largely corresponded with the intimal injury observed in the hypercholesterolaemic rats. These results support the hypothesis that CMV may be one of the factors involved in atherogenesis.


[Glomus tumors of the temporal bone--surgical concept and results]. Between 1985 and 1991, 36 patients with glomus tumors of the temporal bone were operated. More than 60% of the patients presented with an advanced disease (Class C and D) according to the initially discret and slowly progressive clinical signs. Pulsatile tinnitus, hearing loss and paresis of the lower cranial nerves IV to XII were most often found. The management of the patients requires a complete surgical resection with different approaches depending on the extension of the tumor. High resolution CT of the temporal bone and selective angiography of the tumor feeding vessels proved as reliable and necessary imaging tools to determine the size of the tumor and thereby the surgical procedure. Class A tumors (n = 4) were completely resected by an enaural transmeatal approach. Class B tumors (n = 10) were removed completely in all cases by a combined transmeatal-transmastoid approach. A conductive hearing loss in 2 cases and transient facial paresis in one case were observed. Class C tumors (Glomus jugulare tumors, n = 16) required an infratemporal fossa approach type A. A complete resection was achieved in 87.5%. Class D tumors (n = 6) with intracranial extension were managed in a two stage otoneurosurgical procedure. Due to the tumor size and the required surgical procedure a higher incidence of functional lesions (combined hearing loss, vertigo, cranial nerve pareses) was observed. A facial paresis occurred in all cases but was transient in most of them. The results show that functional conservation surgery for glomus tumors of the temporal bone is only possible in Class A and B and some of the Class C tumors. This requires an early diagnosis.


Daily variations of the involvement of beta-receptors in the sympathetic nerve action on glycogenolysis in perfused rat liver. In perfused rat liver perivascular nerve stimulation (7.5 Hz, 20 V, 2 ms, 5 min) at the liver hilus caused an increase in glucose release, a shift of lactate uptake to output and a decrease in arterial, portal and total flow. The influence of the alpha 1-receptor blocker prazosin and the beta-antagonist propranolol on these nerve effects were studied in the isolated rat liver perfused via both the hepatic artery and the portal vein in three experimental series at 9 a.m., at 2 p.m. and at 8 p.m. 1) The nerve stimulation-dependent increase in glucose output was maximal at 9 a.m. (100%), halfmaximal at 2 p.m. (50%) and very low at 8 p.m. (15%). The alterations in arterial, portal and total flow were similar in all three series. 2) At 9 a.m., at 2 p.m. and at 8 p.m. 5 microM arterial plus portal prazosin nearly completely inhibited the metabolic alterations and blocked the reduction of arterial, portal and total flow by 80%. 3) At 9 a.m. 10 microM arterial, portal and arterial plus portal propranolol inhibited the increase in glucose release to less than 25% and the shift of lactate uptake to output to about 50%. At 2 p.m. this inhibitory effect was not observed, neither by selective arterial or portal, nor by simultaneous arterial plus portal addition of 10 microM propranolol. At 8 p.m. the influence of propranolol could not be studied because of the too small control values. The nerve stimulation-dependent reduction of arterial, portal and total flow was not influenced by propranolol, neither at 9 a.m. nor at 2 p.m.(ABSTRACT TRUNCATED AT 250 WORDS)


Macrophages in tissues and in vitro.

Macrophages in tissues and in vitro. Macrophages have specialized functions in different tissue microenvironments such as lymphohaemopoietic organs and the nervous system. Recently, progress has been made in defining cellular and molecular properties of isolated and tissue macrophages in the developing and adult animal.


The mannose receptor and other macrophage lectins. The macrophage expresses a variety of cell surface lectins with activities that support specific functional roles and correspond to various differentiation states characteristic of this cell type. Recently, research has been carried out to investigate the mannose receptor, the advanced glycosylation end products receptor, the mannose-6-phosphate-receptor, the beta-glucan receptor, sialoadhesin and several galactose-specific binding proteins.


Delineation of the phagocyte NADPH oxidase through studies of chronic granulomatous diseases of childhood. The phagocyte NADPH oxidase is a complex system consisting of membrane and cytosolic components that must assemble at the membrane for proper activation. Studies of patients with chronic granulomatous diseases of childhood have enabled the molecular characterization of these components, which has led to studies defining their interaction during NADPH complex assembly. Understanding NADPH oxidase assembly provides an opportunity to develop therapeutics for the regulation of this important reaction of inflammation.


Chemistry and functional role of the invariant chain. The invariant chain has two critical properties that regulate the function of the class II glycoproteins with which it associates. It prevents class II molecules from binding antigenic peptides during the early stages of their assembly and transport, and it contains a signal that drives the assembled class II-invariant chain complex to an endocytic compartment. Dissociation of the invariant chain in this compartment is believed to generate class II molecules capable of binding antigenic peptides.


Characterization of mitochondrial imidazoline-guanidinium receptive sites (IGRS) in liver. Some imidazoline and guanidinium antihypertensive drugs display high affinity for a nonadrenergic membrane protein, the imidazoline-guanidinium receptive site (IGRS), which is insensitive to catecholamine and physically distinct from alpha 2-adrenoceptor. In the present report, we characterized IGRS in human and rabbit liver using [3H]idazoxan as radioligand. By performing subcellular fractionation, we showed a significant increase in [3H]idazoxan binding sites on membrane fractions enriched in cytochrome oxidase activity, a mitochondrial marker. A further enrichment in [3H]idazoxan binding (53-fold with respect to the homogenate) was found in a purified preparation of mitochondrial outer membranes. This localization of IGRS will facilitate the characterization of its functional activity in liver.


Is imidazoline site a unique receptor? A correlation with clonidine-displacing substance activity. Many specific hypotensive drugs acting via the central alpha 2-adrenoceptors were designed based on their imidazoline/guanidine structure for use as antihypertensives. This unique structure, which is missing in the alpha 2-adrenoceptor natural ligands, led to the search for an endogenous, nonadrenergic ligand, and later on, for its putative receptor. Indeed, an endogenous ligand designated the "clonidine displacing substance" (CDS), was isolated and purified from bovine brain, and characterized in various cells. The most intriguing feature of CDS is its hypertensive action upon injection into the rostral ventrolateral medulla and its competition with clonidine. Is CDS a natural agonist which is displaced by clonidine or other hypotensive drugs? Does the unique imidazoline/guanidine structure imply a unique recognition site? Recent studies reported that an imidazoline site, distinct from the alpha 2-adrenoceptor, is abundant in many tissues, and it preferentially recognizes the imidazolino-guanidino type ligands. The physiological role of these sites is still not well defined. In the present study we show that the richest tissue in imidazoline sites is human placenta (1800 +/- 100 fmol/mg protein). The sites are distributed on the cell surface, as observed in studies of binding to intact cytotrophoblasts and cultured trophoblasts originating from human placenta. Binding studies show that the imidazoline site displays a unique pharmacological profile distinct from the alpha 2-adrenoceptor (eg, benzylidenamino-guanidine, Ki = 18.9 +/- 13.8 nmol/L for the imidazoline sites and Ki = 768 +/- 299 nmol/L for the alpha 2-adrenoceptors; guanidopyrol, Ki = 11.2 +/- 6.3 nmol/L for imidazoline sites and Ki = 10100 +/- 1515 nmol/L for the alpha 2-adrenoceptors).(ABSTRACT TRUNCATED AT 250 WORDS)


Unfolding of the tertiary structure of specific tRNA and ribosomal 5S RNA from plants as studied with hydroxyl radicals. Ribosomal 5S RNA is present in all eubacterial and eukaryotic ribosomes. Despite a large amount of experimental data on the primary and secondary structures of these types of molecules, details of their tertiary structure and their precise function in protein biosynthesis are still not known. Recently we have proposed a new model for the tertiary structure of plant 5S rRNA. In this study we applied the Fe(II)-mediated cleavage reaction to test the model. The data presented here provide experimental evidence that in the 5S rRNA molecule only a few nucleotides are buried in the tertiary structure. Similar experiments performed with methionine initiator tRNA gave results which imply the difference in its structure when compared with the X-ray structure of yeast tRNAPhe.


Conformational properties of streptokinase--secondary structure and localization of aromatic amino acids. The conformational properties of streptokinase (Sk) have been assessed by several spectroscopic techniques. A solvent accessibility of about 70% of the 22 Tyr residues was found by u.v. perturbation spectroscopy. Fluorescence spectroscopy indicates also the surface localization of the single Trp 6 residue. Circular dichroism (c.d.), infrared (i.r.), and Raman spectra were analysed in order to estimate the contents of secondary structure elements of Sk. Values in the range of 14-23% alpha-helices, 38-46% beta-structures, 10-30% turns and 12-23% residual structures were found. The characteristics of the c.d. spectrum support the classification of Sk as an alpha + beta protein. Effects of temperature, pH, and denaturants were studied by c.d. spectroscopy, and on spin-labelled Sk, by e.p.r. spectroscopy. Structural effects were induced at temperatures above 40 degrees C, pH values below 3.0 and urea concentrations above 2 M. At temperatures above 70 degrees C, at pH 2.1, and at urea and Gu.HCl concentrations of 7 M and 5 M, respectively, no further structural changes are revealed in the spectra. At temperatures around 50 degrees C, at pH 3.0, and denaturant concentrations of about 1 M Gu.HCl and 1 M to 2 M urea, c.d. effects were observed in the near-u.v. region indicating an increase in the asymmetry for aromatic amino acids in comparison with the structure of Sk in low ionic strength buffers at neutral pH, 20 degrees C and in the absence of denaturants. These effects were most pronounced for the temperature dependence of the c.d. spectra. E.p.r. spectroscopy has shown that loosening of the protein surrounding of the spin label already begins at 1 M urea and that the mobility of the spin label points to a structural change in Sk at 46 degrees C.


[The prognostic significance of inflammatory cells in malignant human soft tissue tumors. Malignancy grading]. These studies are based on an analysis of 160 spindle cell and/or pleomorphic malignant soft tissue tumors of adults and are aimed at providing an answer to the question of whether or not inflammatory cells in tumor tissue are of prognostic importance with regard to overall survival time. In a univariate analysis, granulocytes and lymphocytes did not show any statistical correlation with survival periods, whereas such correlation was revealed with significance to survival periods by plasma cells and mast cells. With multivariate analysis used and presence of necrotic areas included, only mast cell infiltration remained to be significantly correlated with survival time. Also presented in this paper is a proposed scheme of malignancy grading in which two properly established indicators are scored together with "mast cell infiltration", that is "mitosis count" and "necrosis amount". The three grades of malignancy resulting from this grading procedure are shown to be related with high significance to overall survival time.


The influence of density and size of particles on rumination and passage from the reticulo-rumen of sheep. Plastic particles with different densities (0.92, 1.03, 1.22 and 1.44 g/ml) and sizes (1, 10 and 20 mm) were introduced into the rumen of fistulated sheep kept on a roughage diet. The forestomach was emptied 12 and 24 h after the introduction of the particles, and the contents were replaced by the same amount of rumen contents without plastic particles. The proportions of particles which left the reticulo-rumen (RR) during the experimental period were determined by collecting the faeces during the following 5 d. Non-ruminated particles were separated from the dried RR contents and from the faeces. Large particles were ruminated independently of particle size and density within the investigated range. After 12 and 24 h, 59 and 81% respectively of the particles initially introduced were comminuted due to rumination. During the 12 h period about four times as many particles with a density of 1.44 g/ml passed from the RR into the omasum compared with particles with a density of 0.92 or 1.03 g/ml. Three to ten times more 1 mm particles were excreted than originally-large particles (10 and 20 mm). Particles introduced with an original size of 10 or 20 mm were recovered mostly comminuted in the faeces. In a further experiment the rumens of eight sheep were emptied and filled with a buffer solution. Plastic particles (10 g) of each length (1, 5, 10 and 20 mm; all with a density of 1.03 g/ml) were introduced into the ventral rumen. Sedimentation of particles was prevented by gassing the solution in the RR. Of the initially introduced particles, 31.9, 25.4, 12.7 and 1.5% of the 1, 5, 10 and 20 mm long particles respectively left the RR within 4 h. It is concluded that rumination of particles is independent of particle density and size within the tested range. The probability of particles leaving the RR increases with the higher particle density and with the smaller size. If particle sedimentation is prevented in the RR even 10 mm long particles can leave the RR in considerable amounts.


The effect of sucrose supplements on particle-associated carboxymethylcellulase (EC 3.2.1.4) and xylanase (EC 3.2.1.8) activities in cattle given grass-silage-based diet. Carboxymethylcellulase (EC 3.2.1.4; CMCase) and xylanase (EC 3.2.1.8) activities were assayed in rumen fluid and from microbes closely associated either with rumen particulate material or with feed particles incubated in nylon bags in the rumen of cattle. The cattle were fitted with a permanent rumen cannula and a simple 'T'-piece duodenal cannula and were given four diets in a 4 x 4 Latin Square experiment. The basal diet (diet C) consisted of grass silage, barley and rapeseed meal (700, 240 and 60 g/kg total dry matter (DM)) given at the rate of 5.3 kg/d or supplemented with 1.0 kg sucrose/d given twice daily (diet S), twice daily with 0.25 kg sodium bicarbonate/d (diet B) or as a continuous intrarumen infusion (diet I). Giving sucrose supplements decreased CMCase and xylanase activities extracted from microbes associated with rumen particulate material or feed particles incubated in nylon bags as compared with diet C. Supplementation of the sucrose diet with sodium bicarbonate resulted in higher CMCase and xylanase activities than other sucrose diets (S and I). Particle-associated CMCase and xylanase activities were found to be very sensitive in detecting differences in the rumen environment and were related to changes in cell wall digestion. The activities were highly correlated with disappearance of DM and neutral-detergent fibre from nylon bags incubated in the rumen, rumen and total digestion of cell-wall carbohydrates and rumen pool size of cell-wall carbohydrates. It was concluded that the attachment of fibrinolytic enzymes is involved in the depression of fibre digestion. Particle-associated CMCase and xylanase activities were much higher when measured from rumen particulate material than from feed particles incubated in nylon bags.


An electrophysiological and spectroscopic study of the properties and structure of biological calcium channels. Investigations of a model ion channel. The N- and C-terminally protected peptide N-acetyl-Asp-Phe-Ala-Asn-Arg-Val-Leu-Leu-Ser-Leu-Phe-Thr-Ile-Glu-Met-Leu -Leu-Lys-Met-Leu-NH2, closely based on the sequence of the putative S2 membrane spanning helix of domain II of the dihydropyridine receptor calcium channel of the T-system of skeletal muscle, residues 465-486 (Tanabe et al. (1987) Nature 328, 313-318) has been synthesised. Conductance measurements in planar lipid bilayers show that the peptide is capable of inducing the transmembrane passage of calcium and barium ions, in preference to monovalent cations. No anion conductance is observed. 1H-NMR spectroscopy demonstrates that in an amphilic solvent, methanol, the peptide forms highly stable structures characterised by very slow exchange with solvent of peptide N-H protons. Double-quantum filtered phase-sensitive COSY shows that, on the basis of NH-CH alpha scalar coupling constants, most peptide torsion angles are appropriate to an overall alpha-helical conformation; the presence of some alpha-helix is also supported by CD measurements. Most side-chain connectivities have been identified in a DIPSI-TOCSY experiment. This evidence has been used to construct a low-resolution model of the ion-conducting channel of the muscle T-system dihydropyridine receptor from the sequences of the four homologous putative channel-lining stretches. It is characterised by an association of acidic residues at the putative extra-membranous face of the channel, followed by a predominantly hydrophobic band. The next prominent feature of the model is an ordered array of four acidic residues (glutamates 100, 478, 846 and 1164), followed by four lysines (104, 482, 850 and 1168) which may play a gating role.


Identification and purification of the aspartate/glutamate carrier from bovine heart mitochondria. The aspartate/glutamate carrier from bovine heart mitochondria was solubilized with dodecyl-octaoxyethylene ether (C12E8) and purified by chromatography on hydroxyapatite and celite. On SDS gel electrophoresis, the purified aspartate/glutamate carrier consisted of a single protein band with an apparent Mr of 31,500. When reconstituted into liposomes the aspartate/glutamate carrier protein catalyzed an N-ethylmaleimide-sensitive aspartate/aspartate exchange. It was purified 620-fold with a recovery of 17.2% and a protein yield of 0.03% with respect to the mitochondrial extract. The properties of the reconstituted carrier, i.e. requirement for a counteranion, substrate specificity and inhibitor sensitivity, were similar to those of the aspartate/glutamate carrier as characterized in mitochondria.


Lack of effect on the sodium efflux of the microinjection of D-Ins(1,4,5)P3 into ouabain-poisoned barnacle muscle-fibers. A study has been carried out using relatively intact mature muscle fibers from the barnacle Balanus nubilus to see whether D-Ins(1,4,5)P3 stimulates the ouabain-insensitive Na efflux following its microinjection and whether this is accompanied by a contraction of the fiber. Part of the impetus for a study of this type came from the on-going debate between Vergara, Rojas and co-workers and Lea and co-workers, the former group holding the view that skinned barnacle fibers and skeletal fibers in general are responsive to this isomer. The evidence brought forward indicates that the injection of D-Ins(1,4,5)P3 in concentrations in the range of 10(-2) M to 10(-6) M into cannulated unskinned fibers pretreated with ouabain fails to increase the residual efflux, and additionally fails to elicit a contraction. A similar picture emerges with the use of non-hydrolyzable DL-Ins(1,4,5)P3[S]3, analog following its injection in a concentration of 0.5 microM. Higher concentrations of the analog were unavailable for use. This work also involved verification of the idea that an effect might be obtainable in depolarized fibers. However, doubling or tripling K0+ and injection of the isomer or the analog simultaneously failed to support this idea. Since nothing is known as to whether D-Ins(1,4,5)P3 influences the behavior of the Na(+)-Ca2+ exchanger when operating in the reverse mode, experiments were done to check this possibility. ATPNa2 which is though to activate Na(+)-Ca2+ exchange was injected prior to the isomer or the analog but no significant results were obtained. A similar line of reasoning was followed, that of activating the L-type Ca2+ channel by injecting GTPNa2 which in addition is known to activate adenylate cyclase. Again, neither the isomer nor the analog were effective. Thus, the only conclusion possible is that in relatively intact, mature barnacle fibers there is no coupling between the phosphoinositide signalling pathway and two other key systems, viz. the Na(+)-Ca2+ exchanger when operating in the reverse mode and the L-type Ca2+ channel. Equally clear is that for some unknown reason the ouabain-insensitive Na efflux and the contractile apparatus are insensitive to D-Ins(1,4,5)P3[S]3.


Molecular basis for Glanzmann's thrombasthenia (GT) in a compound heterozygote with glycoprotein IIb gene: a proposal for the classification of GT based on the biosynthetic pathway of glycoprotein IIb-IIIa complex. The genetic basis for Glanzmann's thrombasthenia (GT) was elucidated on a compound heterozygote with glycoprotein (GP)IIb gene: an opal mutation at the end of exon 17 (CGA----TGA) results in only a trace amount of GPIIb mRNA, and a splicing mutation at the acceptor site of exon 26 (CAG----GAG) causes an in-frame, exon skipping process from exon 25 to 27. This aberrant transcript encodes a single-chain polypeptide characterized by a 42-amino acid deletion, which includes the proteolytic cleavage site(s) and a unique, proline-rich region at the location corresponding to the carboxyl-terminal of the normal GPIIb alpha-chain. These characteristics are shared by a previously reported defective GPIIb molecule, which is neither assembled with GPIIIa nor transported to the cellular surface. Despite its normal transcription level, expression of the present defective GPIIb molecule was significantly decreased (approximately 6% of the control level). Because the precursor GPIIb molecule is assembled with GPIIIa in the endoplasmic reticulum (ER) and its processing, as well as stability, is dependent on the GPIIIa subunit, the defective GPIIb molecule may be rapidly degraded by the intrinsic quality control system of the ER due to its inability to form a stable heterodimer complex as a consequence of its misfolded structure. Although we did not confirm that the GPIIIa genes of this individual were normal, GPIIIa may be secondarily decreased (approximately 11% of control), because a large part of it could not be complexed, making it vulnerable to proteolysis. To elucidate the molecular basis for GT, we propose here a classification of GT based on the biosynthetic pathway of the GPIIb-IIIa complex.


Molecular epidemiology of Burkitt's lymphoma from South America: differences in breakpoint location and Epstein-Barr virus association from tumors in other world regions. We have previously shown that the endemic (African) and sporadic (North American) forms of Burkitt's lymphoma (BL) differ at a molecular level. We have now extended our studies to the molecular epidemiology of BL in South America, specifically to two climatic regions: temperate (Argentina and Chile) and tropical (Brazil). We have examined the patterns of chromosomal breakpoint locations in 39 tumors with respect to geography and Epstein-Barr virus (EBV) association. The result of these analyses provide further support for the existence of pathogenetically distinct subtypes of BL in different world regions. The majority of breakpoints on chromosome 8 in South American BL (41%) occurred in the immediate flanking region of c-myc, ie, further 5' of the "typical" sporadic breakpoints, in the first exon/intron region, and further 3' of the "typical" endemic breakpoints, which are usually distant from c-myc. However, the distribution of breakpoints on chromosome 14 in tumors from the temperate and tropical regions of South America is similar to that observed in sporadic and endemic tumors. Interestingly, only one tumor with an unrearranged c-myc gene joined to the S mu region of chromosome 14 was observed. This combination was also rarely observed in our earlier series and presumably is either less readily generated by the mechanism that mediates 8;14 translocation or requires other, infrequent genetic changes to provide the necessary selective advantage for lymphomagenesis. The frequency of EBV association in South American BL (51%) is also intermediate with respect to tumors from the United States (30%) and Africa (100%). No correlation with the breakpoint location on chromosome 8 was discernable. Surprisingly, only 54% of tumors with breakpoint outside c-myc were EBV positive. This is in contrast to endemic tumors and suggests that any pathogenetic contribution of EBV is not dependent on breakpoint location, but is more likely to complement additional pathogenetic elements that differ in different world regions.


Identification of DNA rearrangements at the retinoic acid receptor-alpha (RAR-alpha) locus in all patients with acute promyelocytic leukemia (APL) and mapping of APL breakpoints within the RAR-alpha second intron. Italian Cooperative Study Group "GIMEMA". Seventy patients with acute promyelocytic leukemia (APL) were characterized at the DNA level using genomic retinoic acid receptor-alpha (RAR-alpha) probes on Southern blot experiments. Sixty-two cases were defined as M3 according to the French-American-British (FAB) criteria, and eight had a diagnosis of microgranular or variant (M3v) APL. The use of two restriction enzymes and three probes exploring the second intron of the RAR-alpha gene allowed us to detect specific abnormal DNA fragments in every case, with clustering of rearrangements within the 20-kb intronic region between RAR-alpha exons II and III. A more detailed mapping of APL breakpoints was performed in 52 cases in which three EcoRI subregions of the RAR-alpha second intron were analyzed with corresponding probes. Comparison of clinical and hematological features in the three subgroups of patients with distinct RAR-alpha breakpoints did not show significant differences regarding age, peripheral blood (PB) counts, presence of coagulopathy, or FAB classification (M3 v M3v). Interestingly, a significant difference was observed in the M/F ratio of the three subgroups, with a higher incidence of rearrangements at the 5' end of the RAR-alpha second intron in female patients, and more frequent 3' breakpoints in males. The results of this study indicate that a unique genomic alteration consistently occurs on the 17q- derivative of the APL specific t(15;17) aberration. Moreover, the clinical relevance of RAR-alpha gene analysis both at diagnosis and in follow-up studies is further emphasized.


Granulocyte-macrophage colony-stimulating factor activates microtubule-associated protein 2 kinase in neutrophils via a tyrosine kinase-dependent pathway. Receptors of the hematopoietin superfamily, including the granulocyte-macrophage colony-stimulating factor (GM-CSF) receptor, lack a tyrosine kinase domain as well as other sequences indicative of a known signaling mechanism. In this report, we identify the serine/threonine kinase, microtubule-associated protein 2 (MAP2) kinase, as an intermediate in the GM-CSF signal transduction pathway. Treatment of peripheral blood neutrophils or terminally differentiated HL-60 cells with GM-CSF induced a rapid and dose-dependent increase in MAP2 kinase activity. Maximal activity occurred within 5 minutes and the kinetics of the response varied depending on the target cell (prolonged in neutrophils and transient in neutrophilic HL-60 cells). MAP2 kinase activity in these cells correlates with the induction of a 42-Kd tyrosine phosphoprotein. Furthermore, tyrosine phosphorylation is necessary for MAP2 kinase activation since its activity is inhibited by treatment with the tyrosine kinase inhibitor, erbstatin analog. These data suggest that tyrosine phosphorylation is important in GM-CSF-mediated signal transduction and that MAP2 kinase activation may be a central biochemical event involved in its signaling.


Hodgkin's disease.

Hodgkin's disease. The epidemiological features of Hodgkin's disease (HD) suggest that it is a heterogeneous condition which may have different aetiologies in different age groups. The risk factors for the development of HD in young adults suggest that delayed exposure to a common infectious agent may be involved in this age group. Seroepidemiological studies have shown that HD patients have elevated antibody titres to Epstein-Barr virus (EBV) and the elevated titres have been shown to precede the diagnosis of HD. Recent molecular studies provide support for the idea that EBV is involved in the pathogenesis of HD. EBV genomes are consistently found in a proportion of tumour biopsies, the EBV-infected cells are clonal and the EBV genomes have been localized to Reed-Sternberg cells. Furthermore, EBV latent gene products are expressed by the Reed-Sternberg cells. The majority of HD samples from patients aged greater than 50 years and less than 15 years are EBV positive, whereas the minority (less than 15%) of samples from young adults contain detectable EBV DNA. The results suggest that EBV plays a role in HD in children and older adults but that other agents, possibly other viruses, are involved in young adults.


Receptor-mediated endocytosis of galactose and mannose exposing ligands: an electron microscopic study on adult and neonatal cultured rat hepatocytes. We compared the receptor-mediated endocytosis for galactose and mannose exposing ligands in primary cultures of hepatocytes from newborn and adult rats. The endocytic pathway was revealed ultrastructurally using colloidal gold particles coupled to lactosylated bovine serum albumin and invertase. The binding activity on the cell surfaces is observed by keeping the cells at 4 degrees C. For both ligands used, the binding capacity for hepatocytes from adult rats was greater than for neonatal cultured cells. Increasing the temperature to 37 degrees C, we observed that the protein-gold complexes entered the intracellular endocytic organelles. Within 5-15 min, the marker was confined in vesicles close to the cell surface and in the endosome, while after 60 min, the marker is found in lysosome-like compartments. We found that the process of endocytosis is similar for galactose and mannose exposing ligands. The organelles involved in the process of endocytosis in newborn cultured hepatocytes are not different in shape from those of cultured cells of adult rats, but the process of internalization is slower.


Endothelium-dependent relaxation of rabbit middle cerebral artery to a histamine H3-agonist is reduced by inhibitors of nitric oxide and prostacyclin synthesis. 1. The possible involvement of prostanoids and endothelium-derived relaxing factor (EDRF) in the vasodilatation induced by a histamine H3-agonist was examined in the rabbit perfused middle cerebral artery preconstricted with K+ (50 mM). 2. The endothelium-dependent relaxation to (R)-alpha-methylhistamine [(R)-alpha-MeHA] was competitively antagonized by thioperamide (an H3-antagonist) with a pA2 of 9.05, but unaffected by propranolol, atropine, L- and D-sulpiride. This effect was stereoselective since the (S)-isomer was 100 times less potent than the (R)-isomer. 3. Two inhibitors of nitric oxide synthesis, NG-nitro-L-arginine methyl ester (L-NAME) and NG-monomethyl-L-arginine (L-NMMA), inhibited the relaxation induced by (R)-alpha-methylhistamine. The inhibitory effects of 10(-5) M NG-nitro-L-arginine methyl ester and 10(-5) M NG-monomethyl-L-arginine were reversed by equimolar concentrations of L-arginine, but strongly enhanced by 10(-4) M tranylcypromine. Tranylcypromine alone (10(-5) M-10(-4) M) partially reduced the (R)-alpha-methylhistamine-induced relaxation. Both dexamethasone and indomethacin also inhibited this relaxation. 4. The results suggest that the H3-mediated relaxation of the rabbit middle cerebral artery may involve release of both a prostanoid, probably prostacyclin, and endothelium-derived relaxing factor. The relaxant effects of these two endogenous compounds appear to be synergistic.


One way cross tolerance between cromakalim and salbutamol in the uterus of the rat in vivo. 1. Cross tolerance between the potassium (K+) channel opener, cromakalim and the beta 2-adrenoceptor agonist, salbutamol, was investigated in the uterus of the non-pregnant rat in vivo. Uterine sensitivity to salbutamol was similar in both vehicle-treated and cromakalim-tolerant rats. In salbutamol-tolerant rats, uterine responses to cromakalim were markedly decreased compared with saline-infused rats, such that maximum inhibition of uterine contractions was less than 40%. 2. Propranolol treatment and salbutamol tolerance each produced similar reductions in sensitivity of the uterus to salbutamol of approximately 10 fold. The same dose of propranolol did not influence uterine sensitivity to cromakalim, which suggests that the relaxant action of cromakalim is not due to a direct or indirect activation of uterine beta 2-adrenoceptors. 3. Salbutamol produced a marked (11.7 fold) increase in uterine adenosine 3':5'-cyclic monophosphate (cyclic AMP) concentrations measured ex vivo, which was completely inhibited by propranolol pretreatment, but was unaffected by glibenclamide pretreatment. Cromakalim did not increase uterine cyclic AMP concentrations, suggesting that stimulation of adenylate cyclase is not significant in the uterine relaxant action of cromakalim. 4. The lack of propranolol antagonism of cromakalim and of cromakalim-induced changes in uterine cyclic AMP concentrations suggests that the cross tolerance observed between salbutamol and cromakalim may be at the level of K(+)-channels.


Non-competitive inhibition of GABAA responses by a new class of quinolones and non-steroidal anti-inflammatories in dissociated frog sensory neurones. 1. The interaction of a new class of quinolone antimicrobials (new quinolones) and non-steroidal anti-inflammatory agents (NSAIDs) with the GABAA receptor-Cl- channel complex was investigated in frog sensory neurones by use of the internal perfusion and 'concentration clamp' techniques. 2. The new quinolones and the NSAIDs (both 10(-6)-10(-5) M) had little effect on the GABA-induced chloride current (ICI) when applied separately. At a concentration of 10(-4) M the new quinolones, and to a lesser degree the NSAIDs, produced some suppression of the GABA response. 3. The co-administration of new quinolones and some NSAIDs (10(-6)-10(-14) M) resulted in a marked suppression of the GABA response. The size of this inhibition was dependent on the concentration of either the new quinolone or the NSAID tested. The inhibitory potency of new quinolones in combination with 4-biphenylacetic acid (BPAA) was in rank order norfloxacin (NFLX) much greater than enoxacin (ENX) greater than ciprofloxancin (CPFX) much greater than ofloxacin (OFLX), and that of NSAIDs in combination with ENX was BPAA much greater than indomethacin = ketoprofen greater than naproxen greater than ibuprofen greater than pranoprofen. Diclofenac, piroxicam and acetaminophen did not affect GABA responses in the presence of ENX. 4. In the presence of ENX or BPAA, there was a small shift to the right of the concentration-response curve for GABA without any effect on the maximum response. However, the co-administration of these drugs suppressed the maximum of the GABA concentration-response curve, indicating a non-competitive inhibition, for which no voltage-dependency was observed.5. Simultaneous administration of ENX and BPAA also suppressed pentobarbitone (PB)-gated Icl. On the other hand, both PB and phenobarbitone reversed the inhibition of GABA-induced Ic, by coadministration of ENX and BPAA.6. The effect on GABAA responses of co-administration of new quinolones and NSAIDs was not via an interaction with benzodiazepine receptors coupled to the GABAA receptor, since this effect was not reversed by Rol5-1788 or diazepam.7. It is concluded that the co-administration of new quinolones and some of the NSAIDs inhibit GABAergic transmission, and could result in convulsions.


Subtype classification of the presynaptic alpha-adrenoceptors which regulate [3H]-noradrenaline secretion in guinea-pig isolated urethra. 1. The following experiments were carried out to investigate the presence and type of functional presynaptic receptors in adrenergic nerves of the guinea-pig urethra. 2. The urethra from male guinea-pigs was incubated with [3H]-noradrenaline and superfused with Tyrode solution in vitro. The fractional secretion of [3H]-noradrenaline evoked by 300 electrical pulses was measured. 3. The [3H]-noradrenaline secretion was positively frequency-dependent, yielding a half-maximal secretion at 8 +/- 5 Hz. Stimulation was usually applied at 5 Hz. 4. The [3H]-noradrenaline secretion was not altered by noradrenaline (1 or 100 microM), norephedrine (1 microM), isoprenaline (0.1 microM), 5-hydroxytryptamine (10 microM), oxotremorine (10 microM), adenosine (0.2 mM), propranolol (1 microM), atropine (1 microM) or 8-phenyltheophylline (10 microM). 5. The [3H]-noradrenaline secretion was enhanced by clonidine (3 microM), chlorpromazine (10 microM), metitepine (1 microM), 4-aminopyridine (0.5 mM), tetraethylammonium (2 mM), 3-isobutyl-1-methylxanthine (4 mM), 8-bromo cyclic AMP (1 mM) and forskolin (25 microM). 6. The alpha-adrenoceptor antagonists rauwolscine, yohimbine, phentolamine, prazosin and AR-C 239 maximally enhanced the [3H]-noradrenaline secretion to about 300% of control. The partial alpha-adrenoceptor agonist oxymetazoline maximally enhanced the secretion to about 200% of control. The order of apparent EC50 values was rauwolscine less than yohimbine less than phentolamine less than oxymetazoline less than prazosin less than AR-C 239.7. The enhancing effects of yohimbine (1 microM) with tetraethylammonium (2mM), 8-bromo cyclic AMP (1 mM), or forskolin (25,microM) were additive, but not those of yohimbine (1 microM) with prazosin (10 microM), 4-aminopyridine (0.5 mM), or 3-isobutyl-1-methylxanthine (4 mM).8. These results suggest that the [3H]-noradrenaline secretion in the guinea-pig urethra is regulated by presynaptic alpha2A-adrenoceptors which may, in a cyclic AMP-independent manner, be coupled to a 4-aminopyridine-sensitive potassium channel. The secretion is not influenced by compounds acting at beta-adrenoceptors, muscarinic cholinoceptors or adenosine receptors.


Indirect inhibitory effect of succinylcholine on acetylcholine-activated channel activities and its modulation by external Ca2+ in mouse skeletal muscles. 1. The effect of extracellular calcium on single acetylcholine (ACh)-activated channel activities when desensitizing concentrations of succinylcholine (SuCh) were applied to the surrounding endplate membrane was investigated by the cell-attached patch-clamp technique at endplates of single skeletal muscle (flexor digitorum brevis) fibres of adult mice. 2. Bath-applied SuCh (0.1-3 microM, in 2.5 mM Ca2+) increased in a concentration-dependent manner the mean open time of ACh-activated channel currents recorded at membrane potentials which cancelled the SuCh-induced depolarizations. 3. In the presence of 0.5 and 2.5 mM external Ca2+, SuCh (3 microM) applied outside the patch pipette prolonged the mean open time of ACh-activated channel currents in a time-dependent manner (by 45% and 52%, respectively), and simultaneously significantly decreased the single channel conductance (by 14% and 10%, respectively). These SuCh-induced effects did not occur in a nominally Ca(2+)-free extracellular medium. 4. Under the same conditions, SuCh (3 microM) augmented the time-dependent decline in the opening frequency of ACh-activated channel currents obtained in nominally Ca(2+)-free medium. 5. These results suggest that external calcium ions act to modulate nicotinic ACh receptor channel activity, and accelerate desensitization of the receptor.


Characterization of vascular neuropeptide Y receptors. 1. In the present study we compared neuropeptide Y (NPY) and NPY-related analogues for their ability to activate or bind to vascular NPY receptors in four experimental set-ups. Previous results have suggested the existence of different receptor subtypes, Y1 receptors requiring full-length NPY (1-36) or [Pro34]-NPY, and Y2 receptors recognizing also N-terminally truncated forms of NPY but not [Pro34]-NPY. 2. NPY 1-36 and [Pro34]-NPY dose-dependently increased arterial pressure in the anaesthetized rat with a similar magnitude and potency. NPY 2-36 was much less potent than NPY 1-36. NPY 4-36 and NPY 11-36 were inactive even at a dose as high as 10 nmol kg-1. 3. NPY 1-36, [Pro34]-NPY, NPY 2-36 and NPY 5-36 concentration-dependently increased the coronary resistance in the Langendorff preparation of the rat. NPY 1-36 and [Pro34]-NPY were equipotent, while NPY 2-36 and NPY 5-36 were about 7 and 20 times less potent. At 0.3 microM, NPY 11-36, NPY 20-36 and NPY 22-36 induced a slight contraction while NPY 23-36 was inactive. 4. NPY 1-36, [Pro34]-NPY, NPY 2-36, NPY 4-36, NPY 5-36 and NPY 11-36 evoked concentration-dependent contractions in the isolated inferior caval vein of the rat and guinea-pig. [Pro34]-NPY was more potent than NPY 1-36. NPY 2-36 was equipotent with NPY 1-36, while NPY 4-36, NPY 5-36 and NPY 11-36 were approximately 30 times less potent.5. [Pro34]-NPY was equipotent with NPY 1-36 in displacing the '25I-labelled gut hormone peptide([1251]-PYY) from rat aortic smooth muscle cells, while NPY 2-36 and shorter forms of NPY were much less potent or inactive.6. In caval vein smooth muscle cells of the rat, the displacement pattern was more complex than in aortic smooth muscle cells, in that both [Pro34]-NPY and NPY 13-36 effectively displaced the radioligand,albeit none of them completely.7. In conclusion, the NPY-evoked pressor response in the whole rat and coronary vessels seems to be mediated by vascular Y1 receptors and the binding characteristics of the NPY-related peptides in the aortic smooth muscle cells correspond to a population of such receptors. In the caval vein, the profile of the bioactivity and the binding affinity of the NPY-related peptides suggest a mixed population of Y1/Y2 receptors.


Neuropeptide Y (NPY) receptors in HEL cells: comparison of binding and functional parameters for full and partial agonists and a non-peptide antagonist. 1. We have compared the binding and Ca2+ mobilizing properties of various full agonists, partial agonists and a non-peptide antagonist at the neuropeptide Y (NPY) receptor of human erythroleukemia (HEL) cells. 2. [125I]-NPY binding to intact HEL cells was rapid, saturable, of high affinity and with a specificity typical for the Y1-like subtype: NPY, peptide YY (PYY) and [Pro34]-NPY competed for [125I]-NPY binding with high affinity whereas NPY13-36 and NPY18-36 had only low affinity. 3. NPY, PYY and [Pro34]-NPY potently increased intracellular Ca2+ in HEL cells and had equal efficacy. NPY13-36, vasoactive intestinal peptide (VIP) and pancreatic polypeptide (PP) increased intracellular Ca2+ only poorly. 4. Whereas VIP and PP did not significantly affect NPY-stimulated Ca2+ mobilization, NPY13-36 inhibited NPY-stimulated Ca2+ increases and shifted the NPY concentration-response curve to the right without altering its maximal effect. 5. The agonist (pEC50) potencies of the various peptides corresponded well with the affinities of these compounds in the binding assay (pKi), whereas the antagonist potencies (pKb) of the peptide partial agonists and the pA2 value of the non-peptide NPY antagonist (He 90481), calculated from functional data, were lower than the respective affinities determined in the binding studies. 6. A plot of the fractional Ca2+ response vs the fractional receptor occupancy did not reveal any non-linear receptor-effector coupling for NPY or [Pro34]-NPY; a small receptor reserve might exist for PYY. 7. We conclude that the binding and functional properties of HEL cell NPY receptors are very similar. NPY, PYY and [Pro34]NPY are full agonists at these receptors, whereas NPY13-36 is a partial agonist.


Cytochrome oxidase activity in the nucleus of the tractus solitarius of the cat. We studied the cytochrome oxidase (CO) activity in the nucleus of the tractus solitarius (NTS) of normal cats and in animals subjected to unilateral removal of vagal and glossopharyngeal afferents. In normal cats CO activity was higher in the ventrolateral, dorsolateral, interstitial and ventral NTS subnuclei. The dorsal, medial, commissural and gelatinosus subdivisions showed lower levels of CO activity. The peripheral deafferentation up to 47 days did not reduce the CO activity, suggesting an important role for the central inputs in determining the neural activity of the NTS.


Regulation of the anorectic drug recognition site during glucoprivic feeding. The acute effects of 2-deoxy-D-glucose (2-DG)-induced glucoprivic feeding on the anorectic drug recognition site and Na+K(+)-ATPase in the brain were examined in adult rats and in lean and genetically obese mice. The marked hyperglycemia and the induction of feeding caused by the administration of 2-DG to satiated rats and lean mice were associated with significant increases in Na+K(+)-ATPase activity, and in [3H]ouabain binding and [3H]mazindol binding to the anorectic drug recognition site in hypothalamic membranes. Basal and 2-DG-stimulated levels of blood glucose were significantly correlated to the levels of hypothalamic [3H]ouabain (r = + .91, p less than 0.01) and [3H]mazindol (r = + .87, p less than 0.01) binding. A significant correlation (r = .74, p less than 0.05) was also observed between [3H]mazindol binding and [3H]ouabain binding supporting the hypothesis that these hypothalamic binding sites are functionally coupled in their response to circulating glucose. Following the intracerebroventricular (ICV) administration of the diabetogenic drug alloxan, 2-DG did not stimulate feeding or increase [3H]mazindol and [3H]ouabain binding sites in the hypothalamic paraventricular area. Since 2-DG still caused hyperglycemia in alloxan-treated rats, alloxan-induced inactivation of glucoreceptor mechanisms led to an uncoupling of the anorectic drug recognition site from a hypothalamic glucostat. In genetically obese mice (ob/ob), 2-DG also could not induce feeding or increase hypothalamic [3H]ouabain or [3H]mazindol binding, despite a significant hyperglycemic response. In contrast, 2-DG did increase feeding and the binding of [3H]ouabain and [3H]mazindol to the hypothalamus of lean littermates.(ABSTRACT TRUNCATED AT 250 WORDS)