Biophysical properties of a major membrane phospholipid, dielaidoylphosphatidylethanolamine, found in an Escherichia coli fatty acid auxotroph.

Dielaidoylphosphatidylethanolamine, a principal lipid component of membranes of Escherichia coli fatty acid auxotrophs enriched in elaidic acid, has been studied by paramagnetic resonance, fluorescence, and calorimetric methods. EPR measurements with perdeutero-di-tert-butylnitroxide and 2,2,6,6-tetramethyl piperidine-1-oxyl indicate that, when dispersed in aqueous media, this phospholipid undergoes an abrupt order leads to disorder transition at 37.5 degrees C and 36.5 C, respectively. A similar transition temperature is suggested by experiments with 9-doxyl-dimyristoylphosphatidylethanolamine (DEPE). cis- and trans-Parinaric-acid fluorescence polarization measurements indicate that the midpoint of this transition occurs at 34.0 degrees C and 35.5 degrees C, respectively. Differential scanning calorimetry of DEPE revealed a single, sharp endotherm at 38.5 degrees C with increasing temperature; two exotherms of similar magnitude were observed at 36.5 degrees C and 34.5 degrees C upon cooling. This double transition was not observed by any of the other methods. From these results we conclude that the major structural transition at 30-31 degrees C observed previously with 5-, 12-, and 16-doxyl stearate in intact E. coli membranes is due to the DEPE present (Morrisett, J.D., Pownall, H.J., Plumlee, R.T., Smith, L.C., Zehner, Z.E., Esfahani, M., and Wakil, S.J. (1975) J. Biol. Chem. 250, 6969-6976).


Contribution of cyclic adenosine 3':5'-monophosphate to the regulation of bacterial glycogen synthesis in vivo. Effect of carbon source and cyclic adenosine 3':5'-monophosphate on the quantitative relationship between the rate of glycogen synthesis and the cellular concentrations of glucose 6-phosphate and fructose 1,6-diphosphate in Escherichia coli.

When either fructose, glycerol, or succinate served as a sole source of carbon and energy in nitrogen-starved cultures of Escherichia coli W4597(K) the values of the kinetic constants of the equation that expresses the relationship between glycogen synthesis and hexose phosphates were different from the values observed when glucose was the sole source of carbon and energy. Addition of glucose during either exponential growth or nitrogen starvation to a culture using one of the other carbon sources slowed the rate of glycogen synthesis and shifted the values of the constants toward the values observed in cultures using glucose alone. Addition of cyclic AMP (cyclic adenosine 3':5'-monophosphate) during exponential growth of a culture using glucose caused the values of the constants to be shifted toward the values observed in cultures using a carbon source other than glucose. In all of the metabolic conditions studied in this report the adenylate energy charge ((ATP + 1/2 ADP)/(ATP + ADP + AMP)) and the level of the rate-limiting enzyme of glycogen synthesis, ADP-glucose synthetase (glucose 1-phosphate adenylyltransferase, EC 2.7.7.27), were the same. The data presented here indicate that the difference we observed in the quantitative relationship for glycogen synthesis is the result of the different cellular levels of cyclic AMP in the cells using glucose and the cells using one of the other carbon sources. Since cyclic AMP does not affect the velocity of ADP-glucose synthetase in vitro, apparently a change in the cellular level of cyclic AMP causes a shift in the cellular level of a presently unknown (and previously undetected) effector of this enzyme. The shift in the level of this effector evidently alters the response of the enzyme in vivo to the substrate glucose 1-phosphate and the activator fructose 1,6-diphosphate.


Shedding of hyaluronate from the cell surface of Rous sarcoma virus-transformed chondrocytes.

Transformation of cultured chick embryo chondrocytes with Rous sarcoma virus gives rise to increased incorporation of isotopic precursors into hyaluronate and decreased incorporation into chondroitin 6-sulfate. Chemical measurements of these glycosaminoglycans showed corresponding changes. Comparison of the kinetics of production of glycosaminoglycan by normal and Rous sarcoma virus-transformed chondrocytes demonstrated (i) that the rate of accumulation in the medium was similar in both cultures, and (ii) that approximately 50% of total glycosaminoglycan produced by the normal chondrocytes, but only 10% of that from the transformed cells, accumulated in the cell layer. Prelabel-chase experiments indicated that cell surface-associated hyaluronate, as measured by release from the cell layer by trypsin treatment, was shed rapidly into the medium and accounted for all of the hyaluronate which accumulated in the medium. Thus we conclude (i) that accumulation of cell surface-associated glycosaminoglycan is dramatically reduced in Rous sarcoma virus-transformed chondrocytes, and (ii) that hyaluronate produced by the transformed chondrocytes is first deposited in the cell-associated extracellular compartment and then rapidly shed into the medium, rather than being secreted directly into the medium.


Regulation of fatty acid synthetase activity. The 4'-phosphopantetheine hydrolase of rat liver.

The 4'-phosphopantetheine hydrolase of rat liver, partially purified by ammonium sulfate precipitation, catalyzes the hydrolysis of the prosthetic group 4'-phosphopantetheine from the holo-fatty acid synthetase. The two products of the action of this enzyme, 4'-phosphopantetheine and apo-fatty acid synthetase, were isolated by DEAE-cellulose chromatography and by chromatography on a Sepharose epsilon-aminocaproyl pantetheine column, respectively. The resultant apo-fatty acid synthetase was quantitated by immunoprecipitation and it was also converted to the holoprotein with a crude preparation of rat liver 4'-phosphopantetheine transferase. Quantitative determination of the hydrolase reaction product, 4'-phosphopantetheine, by amino acid analysis and microbiological assays confirmed the presence of 1 mol of this compound/mol of holo-fatty acid synthetase.


The role of cyclic AMP in aldosterone production by isolated zona glomerulosa cells.

The role of cyclic AMP in the regulation of aldosterone production by adrenocorticotropic hormone (ACTH), angiotensin II (A II), potassium, and serotonin was examined in collagenase-dispersed adrenal glomerulosa cells. The ability of 8-bromo cyclic AMP and choleragen to stimulate maximum aldosterone production indicated that cyclic AMP could act as second messenger for certain of the aldosterone-stimulating factors. The actions of ACTH and choleragen on aldosterone and cyclic AMP production were correlated in dog and rat cells, and a similar relation was seen during stimulation of rat cells by serotonin. In contrast, A II and potassium did not cause changes in cyclic AMP formation while stimulating aldosterone production. Intracellular and receptor-bound cyclic AMP were increased 3-fold by 10(-7) M ACTH but not by A II. Addition of a phosphodiesterase inhibitor increased the magnitude of the cyclic AMP response to ACTH but did not change the lack of stimulation by A II or potassium. In dog cells, the effects of A II and potassium on aldosterone production were partially additive to those of ACTH, choleragen, and 8-bromo cyclic AMP. In contrast, no additivity was observed between A II and potassium, or between combinations of the cyclic AMP-dependent stimuli. These results indicate that the actions of ACTH on aldosterone secretion are mediated by cyclic AMP formation, whereas A II and potassium stimulate aldosterone production through an independent mechanism. The lack of additivity between steroid responses to A II and potassium suggests that these factors could share a common mode of action on steroidogenesis in zona glomerulosa cells.


Characterization of small cAMP-binding fragments of cAMP-dependent protein kinases.

Monomeric cAMP-binding fragments of molecular mass 16,000 and 14,000 daltons were obtained by Sephadex G-75 chromatography of partially trypsin-hydrolyzed regulatory subunits of cAMP-dependent protein kinase isozymes I and II, respectively. The Stokes radii were 19.1 and 16.4 A, the frictional ratios were 1.15 and 1.03, and the sedimentation coefficients were 1.94 and 1.91 S for the 16,000- and 14,000-dalton fragments, respectively. The 16,000-dalton fragment retained specific cyclic nucleotide binding characteristics of the native protein. The specificity of cyclic nucleotide binding to the 14,000-dalton fragment (cAMP greater than cIMP = 8-bromo-cAMP = 8-oxo-cAMP greater than cUMP = cGMP) differed from that of the native subunit (cAMP = 8-oxo-cAMP greater than 8-bromo-cAMP greater than cIMP greater than cUMP = cGMP). The 14,000-dalton fragment bound nearly 1 mol of cAMP/mol of fragment. The binding exchange rate of cAMP was much faster for the 14,000-dalton fragment than for either of the native regulatory subunits or for the 16,000 dalton fragment. Although hemin inhibited cAMP binding to the native regulatory subunits and to the 16,000 dalton fragment, the molecule did not affect cAMP binding to the 14,000-dalton fragment. Both of the native regulatory subunits and the isolated 16,000- and 14,000-dalton fragments could be covalently labeled with the photoaffinity analog, 8-N3-[32P]cAMP. The 14,000-dalton fragment could not be phosphorylated and neither fragment could recombine with the catalytic subunit to inhibit its activity. The results indicate that the functional entities of the regulatory subunit other than cAMP binding are destroyed by trypsin. The properties of the 16,000-dalton fragment suggest that the intact cAMP-binding site is contained in a small trypsin-resistant "core" of the native regulatory subunit. The properties of the 14,000-dalton fragment imply that part of the binding site of the native regulatory subunit was slighlty modified or lost during preparation of this fragment.


Purification of a reconstitutively active iron-sulfur protein (oxidation factor) from succinate . cytochrome c reductase complex of bovine heart mitochondria.

Oxidation factor, a protein required for electron transfer from succinate to cytochrome c in the mitochondrial respiratory chain, has been purified from isolated succinate . cytochrome c reductase complex. Purification of the protein has been followed by a reconstitution assay in which restoration of ubiquinol . cytochrome c reductase activity is proportional to the amount of oxidation factor added back to depleted reductase complex. The purified protein is a homogeneous polypeptide on acrylamide gel electrophoresis in sodium dodecyl sulfate and migrates with an apparent Mr = 24,500. Purified oxidation factor restores succinate . cytochrome c reductase and ubiquinol . cytochrome c reductase activities to depleted reductase complex. It is not required for succinate dehydrogenase nor for succinate . ubiquinone reductase activities of the reconstituted reductase complex. Oxidation factor co-electrophoreses with the iron-sulfur protein polypeptide of ubiquinol . cytochrome c reductase complex. The purified protein contains 56 nmol of nonheme iron and 36 nmol of acid-labile sulfide/mg of protein and possesses an EPR spectrum with the characteristic "g = 1.90" signal identical to that of the iron-sulfur protein of the cytochrome b . c1 complex. In addition, the optimal conditions for extraction of oxidation factor, including reduction with hydrosulfite and treatment of the b . c1 complex with antimycin, are identical to those which facilitate extraction of the iron-sulfur protein from the b . c1 complex. These results indicate that oxidation factor is a reconstitutively active form of the iron-sulfur protein of the cytochrome b . c1 complex first discovered by Rieske and co-workers (Rieske, J.S., Maclennan, D.H., and Coleman, R. (1964) Biochem. Biophys. Res. Commun. 15, 338-344) and thus demonstrate that this iron-sulfur protein is required for electron transfer from ubiquinol to cytochrome c in the mitochondrial respiratory chain.


Pitfalls to avoid: accidental intravenous injection of Ethiodol.

Direct intravenous injection of Ethiodol occurred at lymphography. Early fluoroscopic or radiographic film evaluation, and careful monitoring of injection rates, would help to avoid this complication.


Prostaglandin E2 and cyclic AMP levels in human breast tumors.

Prostaglandin E2 (PGE2) and cyclic adenosine-3', 5'-monophosphate (cAMP) concentrations were measured in human benign and malignant breast tumors by radioimmunoassay. Two groups were found among the malignant tissues, one with high PGE2 (M = 65.89 pg/mg) and high cAMP (M = 0.704 pmole/mg) concentrations and one with low concentrations (M = 9.24 pg/mg and M = 0.299 pmole/mg, respectively). The low PGE2 levels in the malignant tumors did not differ significantly from the levels found in benign tumors (M = 8.06 pg/mg). cAMP levels were positively and highly correlated (r = +0.81) with PGE2 levels. No bone osteolysis could be discovered in any of the patients a few weeks after mastectomy operation, but PGE2 analysis of breast tumors may have prognostic value for the future.


Light and electron microscopic investigations of nasopharyngeal carcinomas with regard to the viral etiology of these tumors.

Five carcinomata of the nasopharynx (four lymphoepithelial carcinomata of the Regaud type and one squamous cell carcinoma) were examined light and electron microscopically. In addition to the familiar histological and cytological features of these tumors, and because of an increased antibody titer against Epstein-Barr virus in all five patients, all those cytoplasmic and nuclear inclusions were examined which could be interpreted as indicative of a virus contact. The following structures were found: 1. Particles and microtubules which correspond in diameter, shape, and location to Corona viruses. 2. Particles surrounded by a double membrane and resembling in form and diameter Oncorna viruses. 3. Tubulo-reticular, coil-shaped cytoplasmic inclusions interpreted as an unspecific reaction of the host cell to viral attack. 4. Spherical nuclear bodies, which are frequently observed in tumors and in viral infections. 5. Intranuclear particles which correspond in diameter, structure, and distribution to viruses of the herpes type such as have been described in cell cultures of Burkitt lymphoma and nasopharyngeal carcinoma. The fifth group particularly was discussed in detail with regard to differentiation between those particles and other structures which could simulate a virus structure. Together with the appearance of increased ribosomes and of particular chromatin distribution within the tumor cell nuclei, the particles we discussed have been interpreted as morphological indications of a virus etiology of the examined tumors.


Changes in mouse skin cyclic nucleotides during chemical carcinogenesis and tumor response to treatment with BCG, L-Dopa and cyclic DBAMP.

During chemical carcinogenesis by 9, 10-dimethyl 1,2-benzanthracene the cyclic AMP and GMP content was measured in the skin of mice. The tissue of the developing skin tumor is characterized by an elevated cyclic GMP- and a lowered cyclic AMP level. Consequently the quotient of cyclic AMP and GMP is greatly lowered and is discussed as a crucial factor of cell derangement. Complete regression of the tumor is to be achieved by increasing the cyclic AMP levels by means of cyclic DBAMP and L-Dopa in 20% of the experimental animals. Additional stimulation by BCG of the unspecific immune defence has a favourable effect on tumor regression (about 50%). In a different type of tumor (solid Ehrlich carcinoma) it was possible after pretreatment of the mice with L-Dopa, cyclic DBAMP and BCG and after treatment following tumor injection to prevent tumor development in 67% of the animals.


Establishment and characterization of C-type RNA virus-producing cell lines from radiation-induced murine osteosarcomas.

Eight cell lines were established from murine osteosarcomas induced in vivo with the radionuclides 224Ra and 227Th. They have been compared by light and electron microscopy, by karyology, and by their growth properties. The morphology, the growth pattern, and the ability to induce tumors in mice indicate that five of them are tumor cell lines. Chromosome studies demonstrated that the five cell lines have marker chromosomes. The other cell lines only showed some criteria generally used to score for transformation of fibroblasts and they may be derived from stromal cells. All cell lines release virus particles in the culture fluid which have the typical properties of RNA tumor viruses. They possess C-type morphology, a density of 1.16--1.18 g/cm3, a 60--70 S RNA, a RNA dependent DNA polymerase and they induce syncytia in rat XC cells. The possible significance of these virus particles in radiation osteosarcomagenesis is discussed.


Sodium-potassium adenosine triphosphatase activity of human lymphocyte membrane vesicles: kinetic parameters, substrate specificity, and effects of phytohemagglutinin.

We have prepared human blood lymphocyte membrane vesicles of high purity in sufficient quantity for detailed enzyme analysis. This was made possible by the use of plateletpheresis residues, which contain human lymphocytes in amounts equivalent to thousands of milliliters of blood. The substrate specificity and the kinetics of the cofactor and substrate requirements of the human lymphocyte membrane Na+, K+-ATPase activity were characterized. The Na+, K+-ATPase did not hydrolyze ADP, AMP, ITP, UTP, GTP or TTP. The mean ATPase stimulated by optimal concentrations of Na+ and K+ (Na+, K+-ATPase) was 1.5 nmol of P(i) hydrolyzed, microgram protein-1, 30 min-1 (range 0.9-2.1). This activity was completely inhibited by the cardiac glycoside, ouabain. The K(m) for K+ was approximately 1.0 mM and the K(m) for Na+ was approximately 15 mM. Active Na+ and K+ transport and ouabain-sensitive ATP production increase when lymphocytes are stimulated by PHA. Na+, K+-ATPase activity must increase also to transduce energy for the transport of Na+ and K+. Some studies have reported that PHA stimulates the lymphocyte membrane ATPase directly. We did not observe stimulation of the membrane Na+, K+-ATPase when either lymphocytes or lymphocyte membranes were treated with mitogenic concentrations of PHA. Moreover, PHA did not enhance the reaction velocity of the Na+, K+-ATPase when studied at the K(m) for ATP, Na+, K+ OR Mg++, indicating that it does not alter the affinity of the enzyme for its substrate or cofactors. Thus, our data indicate that the increase in ATPase activity does not occur as a direct result of PHA action on the cell membrane.


Plasma membrane phosphoproteins in normal and Rous sarcoma virus transformed chick embryo fibroblasts: characterization by in vitro phosphorylation.

Plasma membranes isolated from normal and RSV transformed chick embryo fibroblasts were phosphorylated in vitro using endogenous protein kinase and ATP (gamma32P) and the labeled phosphoproteins were analyzed by SDS-PAGE. A number of protein phosphorylation changes were observed following transformation, however in most cases they were relatively small quantitative differences. The four major changes were in proteins of 47,000, 58,000, 75,000 and 135,000 daltons. Decreased phosphorylation of the 47,000 dalton polypeptide was found in transformed cell membranes but this alteration was shown to be due to differences in cell growth rather than transformation. Increase phosphorylation of the 75,000 dalton protein was at least partially related to virus infection. However, increased phosphorylation of the 58,000 and 135,000 dalton polypeptides were entirely transformation specific.


Long acting cAMP analogues enhance sulfate incorporation into matrix proteoglycans and suppress cell division of fetal rat chondrocytes in monolayer culture.

The relationship between replication and the synthesis of matrix sulfated proteoglycans was investigated with fetal rat chondrocytes grown in monolayer culture. The effect of N6 O2' dibutyryl adenosine 3', 5' cyclic monophosphate (DBcAMP), adenosine 3', 5' cyclic monophosphate (cAMP), 8 Bromo adenosine 3', 5' cyclic monophosphate (8 Br-cAMP), sodium butyrate and hydroxyurea was examined. Between 0.05 and 0.5 mM DBcAMP, a dose related inhibition of cell division and stimulation of [35SO=/4] incorporation into matrix proteoglycans was demonstrated. At the higher concentrations of DBcAMP, cell division was completely inhibited and the enhancement of [35SO=/4] incorporation into matrix proteoglycans ranged between 40 and 120% (P less than 0.01). Utilizing 14C-glucosamine and photometric determination of proteoglycans with Alcian Blue, it was demonstrated that the increase in sulfate incorporation reflected enhanced accumulation of extracellular matrix. The effects of DBcAMP were mimicked by 8 Br-cAMP, suggesting they were mediated by the adenylyl cyclase system. cAMP (0.05-0.5 mM), sodium butyrate (0.1-0.5 mM) and hydroxyurea (0.5-5 mM) partially or fully inhibited cell division, but either failed or only slightly enhanced sulfate incorporation. The enhanced sulfated proteoglycan deposition promoted by DBcAMP began 8 to 12 hours after serum stimulation, its onset occurred prior to thymidine incorporation and the effect persisted for 28 hours. Determination of cell volume demonstrated an increase in size of DBcAMP treated chondrocytes between 8 to 12 hours, coincident with the onset of increased sulfate incorporation. These results are consistent with a model where matrix sulfated proteoglycan deposition by chondrocytes is mediated by intracellular cAMP levels and occurs in the G1 phase of the cell cycle.


Effect of hypothalamic extract and other factors on release of adrenocorticotropin from and adenosine 3',5'-monophosphate levels in dispersed nonpituitary tumor cells.

Tumor tissues obtained from two patients with the ectopic ACTH syndrome caused by medullary carcinoma of the thyroid and malignant epithelial thymoma were dispersed by tryptic digestion and mechanical agitation. Using the isolated cells, the effects of various agents on ACTH secretion and intracellular cAMP concentrations were studied. Addition of rat median eminence extract significantly stimulated ACTH secretion and increased levels of intracellular cAMP in both cell preparations, and a dose-response relationship appeared to exist between the dose of rat median eminence extract added and either ACTH secretion or intracellular cAMP formation in the thymic tumor cells. High concentrations of calcium also produced a marked ACTH secretion in both cases. In the thymic tumor cells, norepinephrine, serotonin, and TRH were found to be effective in increasing ACTH secretion and intracellular cAMP levels, whereas biogenic amines, hypothalamic hormones, and gastrointestinal hormones did not affect hormone secretion in the thyroid tumor cells. These results suggest that a corticortropin-releasing factor-like substance(s), as yet unspecified, may play some role in stimulating ectopic ACTH secretion by certain tumors, that both intracellular cAMP and Ca++ may be involved in ectopic hormone secretion, and that the inappropriate hormonal secretory responses of some tumors to a variety of stimuli might be mediated by altered membrane receptors of the neoplastic cells.


An attenuated form of congenital virilizing adrenal hyperplasia.

The families of two patients with hirsutism and oligomenorrhea were studied with iv ACTH (Cortrosyn) stimulation. The parents responded with a combined incremental rise of progesterone and 17-hydroxyprogesterone greater than 6.5 ng/dl.min, a response seen in the heterozygote parents of patients with congenital virilizing adrenal hyperplasia (CVAH). One sibling in one family responded as do heterozygous subjects, while two other siblings responded normally. The abnormal response of the two pairs of parents and the sibling along with the pattern of steroid secretion observed in the propositi suggest that the so-called adult-onset CVAH is a mild form of the homozygous state of CVAH.


Plasma levels of adrenocorticotropin and cortisol in women receiving oral contraceptive steroid treatment.

The secretion rate and plasma concentration of the adrenocortical steroid cortisol is modified in subjects treated with estrogenic and/or progestational steroids. The effects of contraceptive steroids on the secretion of ACTH are poorly documented, however, In the current investigation, we found that concentrations of ACTH and cortisol in plasma obtained at 0800--0900 h from a group of women with normal cyclic menses (n = 4) ranged from 78--120 pg/ml and 77--137 ng/ml, respectively. Although significant cyclic changes in the plasma levels of LH, FSH, 17 beta-estradiol, and progesterone occurred during the ovarian cycle, no obvious cyclic fluctuations in plasma levels of ACTH or cortisol were observed. In women treated with Norinyl 1 + 80 (1.0 mg norethindrone plus 0.08 mg mestranol), plasma concentrations of LH, FSH, 17 beta-estradiol, and progesterone were significantly lower (P less than 0.001) than plasma levels of these hormones in normal women during the ovarian cycle. The mean daily plasma concentrations of ACTH were significantly lower (P less than 0.001), whereas plasma cortisol levels were significantly higher (P less than 0.001) in women treated with oral contraceptive steroids compared to the levels of these hormones in the untreated ovulatory women.


Adrenocorticotropin-dependent virilizing paraovarian tumors in Nelson's syndrome.

A 35-yr-old woman with Nelson's syndrome presented with amenorrhea and virilization. Serum testosterone (T) concentration was 605 ng/dl and fell to 33 ng/dl when dexamethasone was administered. The MCR of T fell from 1383 to 991 liters/day and the T production rate decreased by 96%. With administration of synthetic ACTH, T concentration rose to 338 ng/dl. Plasma ACTH concentration paralleled T during repeated suppression testing, suggesting that T secretion was dependent on ACTH hypersecretion. Preoperative and intraoperative ovarian vein catheterization suggested that the predominant source of androgen production was from the right ovarian vein. Laporatomy revealed multiple paraovarian tumors in the right mesosalpinyx and mesovarium. Incubation of tumor slices and ovarian tissue with [3H]pregnenolone and [14C]17-hydroxyprogesterone demonstrated conversion of both precursors to T by the tumor and confirmed that the tumors were the source of androgen excess. The microscopic appearance of the tumors closely resembled the morphology of testicular and paratesticular tumors of men with congenital adrenal hyperplasia and Nelson's syndrome. The analogous dependency of the tumors on ACTH hypersecretion in men with paratesticular tumors and in this woman with paraovarian tumors suggests that the tumors may arise in both males and females from a common steroid-secreting cell of adrenogenital origin.


ACTH, beta-LPH and beta-endorphin in pituitary adenomas of the patients with Cushing's disease: activation of beta-LPH conversion to beta-endorphin.

ACTH, beta-lipotropin (beta-LPH) and beta-endorphin concentrations were determined in pituitary adenomas of the patients with Cushing's disease. Immunoreactive ACTH and beta-endorphin were present in high concentrations and essentially equimolar amounts in pituitary adenomas. beta-LPH conversion to beta-endorphin was activated in pituitaries associated with ACTH/beta-LPH producing adenomas. Immunoreactive ACTH and beta-endorphin concentrations were markedly suppressed in the surrounding tissues.


Altered metabolism (in vivo and in vitro) of plasma lipoproteins after selective chemical modification of lysine residues of the apoproteins.

Chemical modification of lysine residues by acetoacetylation of the apoproteins of iodinated canine and human low density lipoproteins (LDL) and canine high density lipoproteins (HDL) resulted in a marked acceleration in the rate of removal of these lipoproteins from the plasma after intravenous injection into dogs. Clearance of the lipoproteins from the plasma correlated with their rapid appearance in the liver. Acetoacetylated canine (125)I-LDL (30-60% of the lysine residues modified) were essentially completely removed from the plasma within an hour, and > 75% of the activity cleared within 5 min. Reversal of the acetoacetylation of the lysine residues of the LDL restored to these lipoproteins a rate of clearance essentially identical to that of control LDL. Identical results were obtained with modified human LDL injected into dogs. At 10 min, when congruent with 90% of the acetoacetylated human (125)I-LDL had been removed from the plasma, 90% of the total injected activity could be accounted for in the liver. Furthermore, it was possible to demonstrate an enhancement in uptake and degradation of acetoacetylated LDL by canine peritoneal macrophages in vitro. The mechanism(s) responsible for the enhanced removal of the LDL and HDL in vivo and in vitro remains to be determined. By contrast, however, acetoacetylation of canine (125)I-apoE HDL(c) did not accelerate their rate of removal from the plasma but, in fact, retarded their clearance. Control (native) apoE HDL(c) were removed from the plasma (64% within 20 min) and rapidly appeared in the liver (39% at 20 min). At the same time point, only 45% of the acetoacetylated apoE HDL(c) were cleared from the plasma and <10% appeared in the liver. Acetoacetylation of the apoE HDL(c) did not enhance their uptake or degradation by macrophages. The rapid clearance from the plasma of the native apoE HDL(c) in normal and hypercholesterolemic dogs suggests that the liver may be a normal site for the removal of the cholesteryl ester-rich apoE HDL(c). The retardation in removal after acetoacetylation of apoE HDL(c) indicates that the uptake process may be mediated by a lysine-dependent recognition system.


Regulation of sodium and potassium transport in phytohemagglutinin-stimulated human blood lymphocytes.

Phytohemagglutinin (PHA) or concanavalin A treatment of lymphocytes causes an increase in membrane permeability so that the leak rates of Na and K increase 1.5- to 2-fold. Active Na and K transport increase proportionately in response to the increased membrane permeability. We have examined the role of lymphocyte Na concentration in sustaining the increased Na and K transport observed after PHA treatment. Cell Na concentration increases from 14.8 to 20.5 mmol/liter cell water in PHA-treated lymphocytes (P < 0.001). Four lines of evidence suggest that the 5-6 mmol/liter cell water increase in lymphocyte Na accounts for the increase in active Na and K transport in mitogen-treated lymphocytes. First, PHA does not increase directly the maximal Na, K-ATPase activity of isolated lymphocyte membrane vesicles. Second, when the Na concentration is increased by 6 mmol/liter cell water in unstimulated lymphocytes, Na and K transport increase nearly twofold. Third, the cell Na concentration (15 mmol/liter cell water) is near the K(m) for Na activation of the Na, K-ATPase in lymphocyte membranes. The ATPase activity thus, is capable of increasing as the cell Na rises above normal. Fourth, if lymphocytes are incubated in a medium containing a low Na concentration, K transport does not maintain the internal K concentration and the fall in cell K is accentuated in PHA-treated lymphocytes. These studies indicate that the adaptive acceleration of Na and K transport in mitogen-treated lymphocytes is mediated by a small increase in cell Na.


A simple biological method for detecting streptococcal nicotinamide adenine dinucleotide glycohydrolase.

A biological method for detecting streptococcal nicotinamide adenine dinucleotide glycohydrolase (NADG) is presented, based on its ability to inhibit the growth of Haemophilus parainfluenzae. Three hundred clinical isolates of beta-haemolytic streptococci were tested. All isolates producing NADG belonged to Lancefield's group A, C, or G.


Bone metastases in children presenting with renal tumours.

A survey of bone metastases in 132 children presenting with renal tumours has been carried out using material from the Manchester (England) Children's Tumour Registry during the period 1954 to 1976. Seven such cases were found. Only one of the 117 Wilms' tumours developed bone metastases. The remainder included carcinoma (1/4), 'bone metastasising renal tumour of childhood' (2/3), atypical tumours (2/2), and lymphosarcoma (1/1). Bone metastases from true nephroblastoma seem to be an exceptional finding.


Angiofibromas in tuberous sclerosis: a light and electron microscopic study.

Angiofibromas from two patients with tuberous sclerosis were studied by light and electron microscopy. Light microscopy revealed that these tumor-like nodules (which in the past have been called adenoma sebaceum) were made up of dilated capillaries, venules and arterioles embedded in connective tissue. At the ultrastructural level the arterioles embedded in connective tissue. At the ultrastructural level the endothelium of these vessels showed large numbers of microvilli on their luminal surface. The stroma contained many banded structures (so-called fibrous long spacing collagen). Myofibroblasts recently described in juvenile nasopharyngeal angiofibromas were not found in these angiofibromas of tuberous sclerosis.


The relative caries-inhibiting effects of a stannous fluoride dentifrice in a silica gel base.

An unsupervised toothbrushing study involving 1,339 children from 5 to 13 years of age conducted for three years compared two stannous fluoride dentifrices, one in a calcium pyrophosphate base and the other in a silica gel base, with a nonfluoride control dentifrice. The test dentifrice, stannous fluoride in a silica gel base, reduced caries to a significant extent when compared with the nonfluoride control dentifrice.The percentage of reductions ranged from 15% to 25% for whole mouth and interproximal surface indexes. There was no significant difference between the two fluoride dentifrices.


Simultaneous development of autoimmune disease and malignancy in two elderly patients.

Two elderly patients who presented with symptoms of "systemic rheumatic or autoimmune disease" had high serum titers of fluorescent antinuclear antibody (ANA). Signs of malignancy were evident at the time of the first examination. T-cell dysfunction in the aged may permit the simultaneous development of autoimmunity and neoplasia. When rheumatic symptoms with a high titer of fluorescent ANA develop in elderly patients, an exhaustive search for malignancy should be considered.


Studies on the origin of the methylsulfonyl-containing metabolites from propachlor.

Metabolites in which the chlorine from propachlor has been replaced by a cysteine group or a methylsulfonyl group [-S(O2) CH3] are present in the urine of rats dosed orally with propachlor. In the present study, urine from rats given single oral doses of 35S-labeled cysteine conjugate of propachlor contained metabolites having the methylsulfonyl groups labeled with 35S. No metabolites containing 14C-labeled methylsulfonyl groups were isolated from urine of rats given single oral doses of the cysteine conjugate of propachlor in which the cysteine group was uniformly labeled with 14C. These findings show that the cysteine conjugate of propachlor is the source of sulfur in the methylsulfonyl-containing metabolites. Therefore, we suggest that a C-S lyase present in the animal cleaves the cysteine conjugate of propachlor and thus allows further metabolism of the sulfur to a methylsulfonyl moiety.


Ion movements in the nervous system of the American cockroach, Periplaneta americana (L.), influenced by toxaphene.

The effect of toxaphene upon ion fluxes of the central nervous system (CNS) of the cockroach, Periplaneta americana (L.), was studied in vitro. The CNS, divided into three sections (brain, thoracic, and abdominal), was exposed to 10(-5) M toxaphene in saline containing 24Na+, 42K+, 36CL-, or 45Ca++. Uptake and efflux of these ion were evaluated. Toxaphene was responsible for significant increases in the internal levels of 42K+ and 45Ca++, but had little effect upon 24Na+ or 36Cl- movements. The resulting concentration changes of the ions may be involved in the neural activity observed with toxaphene poisoning. Of the three CNS sections, significant changes were more numerous and occurred earlier in abdominal sections.


Adenosine triphosphatase activities in brain, kidney and liver of mice treated with toxaphene.

The sensitivity of Na+-K+ and Mg++ Adenosine Triphosphatases (ATPases) in mouse tissues to toxaphene, a highly chlorinated camphene, was determined both in vivo and in vitro. The brain and kidney Na+-K+ ATPase activities were significantly inhibited in vitro by toxaphene. Interestingly, the inhibition was not significantly increased with an increase in the concentration of toxaphene. The oligomycin-sensitive (mitochondrial Mg++ ATPase activities in mouse brain, kidney and liver fractions were significantly inhibited by toxaphene in a concentration-dependent fashion. The oligomycin-insensitive Mg++ ATPase in all tissues examined was also inhibited but less sensitive to toxaphene than mitochondrial Mg++ ATPase. In contrast to in vitro response, the brain ATPases were not altered in mice fed toxaphene by oral intubation for three days. The renal and hepatic ATPase activities were significantly decreased in toxaphene treated mice with the oligomycin-insensitive Mg++ ATPase activity being only slightly altered.


Deformities of the hand and wrist with ulnar deficiency.

Deformities of the hand and wrist in 26 upper limbs of 20 patients with ulnar deficiency were studied. In eight limbs the ulna was totally absent. In the remainder it was severely or moderately hypoplastic. Abnormalities of the digits were present in all affected limbs. There was ectrodactyly in 25 hands. The thumb was hypoplastic in 21 hands and totally absent in five. Digital deformities also included syndactyly (14 hands) and delta phalanx (six hands). Early syndactyly release, pollicization, and rotational osteotomy of the metacarpals improved hand function in many of these patients. In only three of 26 limbs was there more than 20 degrees of fixed ulnar deviation at the wrist. In 16 patients the distal ulnar "anlage" was not removed. None of these patients developed ulnar deviation of the wrist or progressive deformity at the wrist during growth. Resection of the distal ulnar "anlage" does not appear to be warranted, unless there is progressive deformity of the wrist or elbow or increased bowing of the radius.


Multiple digital glomus tumors.

In 28 patients with glomus tumors of the digits, 21 were relieved of symptoms by excision of a solitary tumor. Seven other patients had multiple tumors, four of which were the cause of recurrent symptoms. Three patients had multiple tumors--five in one patient, four in another, and two in a third, which had been found at the time of the first operation. A 25% incidence of multiple tumors and a 15% incidence of recurrent symptoms indicates that a careful search should be made for additional tumors.


Sero-epidemiology of type--2 (genital) herpes simplex virus infection in Ibadan.

Herpes Type 1 (HT-1) virus infection is prevalent in Ibadan, Nigeria. In a study to determine the prevalence of Herpes Type-2 (HT-2) virus infection in this environment by the complement fixation (CF) test, it was shown that antibodies to the virus were not found below the age of 14 years, after which there was a gradual rise from the 16-25-year age group to a peak in the 26-35-year age group. It was suggested that the pattern of distribution of HT-2 antibodies is compatible with the sexual habits of the different age groups, and is in support of the venereal mode of transmission of the virus.


Suggestive evidence for receptors for morphine and methionine-enkephalin on normal human blood T lymphocytes.

This study reports the in vitro influence of morphine, dextromoramide, levomoramide, and methionine-enkephalin upon normal human T blood lymphocytes by using the active and total rosette tests. Morphine and dextromoramide inhibited the percentage of active T rosettes. This effect was completely reversed in the presence of naloxone, their specific antagonist. The specificity was further demonstrated by the absence of the effect of levomoramide, the inactive enantiomere, upon the rosette system. Methionine-enkephalin increased the percentage of active T rosettes. This effect was specifically inhibited by naloxone. These observations suggest that normal human blood T lymphocytes bear surface receptor-like structures for morphine, dextromoramide, and methionine-enkephalin. Such findings may provide a link between the central nervous system and the immune system.


Macrophage restriction of Marek's disease virus replication and lymphoma cell proliferation.

Macrophages are shown to restrict the replication of Marek's disease virus (MDV) and isotope uptake by spleen cells from chickens bearing Marek's disease (MD) tumors. The titer of virus from duck embryo fibroblasts (DEF) co-cultivation with MDV-spleen cells pretreated to deplete marcophages was 4- to 18-fold higher than with untreated cells. Treated MDV-spleen cells increased isotope uptake by 2-fold. These restrictive activities are attributable to macrophage regulation of cell proliferation.


Colloidal silica--aluminum modified--PVP density gradient centrifugation: centrifuge tube wall cell adherence, aggregation, separation properties and comparison to BSA and Ficoll.

A method is described for the inexpensive and easy preparation of colloidal silica--aluminum modified--polyvinylpyrrolidone (CS-AM-PVP) density gradient centrifugation medium. Using density gradient centrifugation, several cell separation and biochemical characteristics were studied: centrifuge tube wall cell adherence, mouse spleen cell density distribution, rebanding properties, mitogen response and cell aggregation. Cell adherence to the centrifuge tube wall using density gradients of CS-AM-PVP was compared with density gradients of bovine serum albumin and Ficoll. Few cells adhered to the centrifuge tube wall when CS-AM-PVP was used as a gradient medium; whereas, significant cell adherence to the centrifuge tube cell wall occurred when bovine serum albumin or Ficoll was used as a gradient medium. The CS-AM-PVP gradient medium did not inhibit the response to mitogens of mouse spleen cells which had been separated into density subpopulations in a discontinuous CS-AM-PVP density gradient, caused a minimum amount of cell aggregation, and was found to be non-toxic.


Comparison of activation and blocking procedures in the use of defined antigen substrate spheres (DASS) for quantitative and visual assessment of conjugate activity.

The use of Sepharose beads in the defined antigen substrate sphere (DASS) immunofluorescent system has been restricted by the need for sophisticated equipment for fluorescence quantitation since reliable subjective reading is not possible. This paper describes a method for visual assessment of results in this system. Antigen is coupled to sodium metaperiodate activated. Cross-linked Sepharose 4B beads and blocked with rhodamine-BSA prior to reduction. This procedure eliminates the use of the hazardous cyanogen bromide activation process and is potentially suitable for use in both direct systems for evaluation of specificity and potency of fluorescent conjugates and in indirect systems for serological screening of sera for specific antibody.


Detection of antibodies to Sendai virus by enzyme-linked immunosorbent assay (ELISA).

An enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies to Sendai virus, a paramyxovirus, is described. The assay was found to be about 20-fold more sensitive than the hemagglutination inhibition assay. Differentiation between virus specific IgG and IgM is possible. The test appears to be especially useful in the study of early events in antibody formation in vivo as well as in vitro.


Comparative ultrastructure and cytochemistry of epidermal responses to tape stripping, ethanol and vitamin A acid in hairless mice.

The responses of hairless mouse epidermis to tape stripping, ethanol and vitamin A acid were compared using electron microscopy and cytochemistry. Stripping the skin 4 times with cellophane tape removed most of the stratum corneum and caused the development of enlarged intercellular spaces and dense intramitochondrial inclusions. These changes began within an hour, reached a maximum by 24 hr, and subsided by about 4 days. There was also some accumulation of glycogen and the development of occasional basal lamina breaks and a few intracellular lipid droplets by 24 hr. Topically applied ethanol (95%) produced a similar response, though less damaging. In addition, ethanol induced the formation of numerous lipid droplets in the cytoplasm of keratinocytes by 24 hr and often caused sloughing of the stratum corneum. Vitamin A acid (3%) was dissolved in propylene glycol for application to the skin. We found the application of propylene glycol alone to produce no epidermal changes. Vitamin A acid produced mild damaging effects with small intercellular spaces and intramitochondrial inclusions, but no lipid droplets or glycogen were detected. Vitamin A acid also caused dramatic inhibition of keratinization by 24 hr. Intramitochondrial inclusions were digested by protease, indicating a protein component and they also stained with silver methenamine, indicating the presence of glycogen or glycoprotein.


The effect of fasting on the adrenergic receptor activity of human adipocytes.

To assess the influence of fasting of the alpha- and beta-adrenergic receptor activities of human adipocytes, subcutaneous adipose tissue was obtained from normal obese subjects in the fed state and after 3 days of fasting. Isolated cells prepared from the adipose tissue were incubated with epinephrine, 10(-6)M, and in the presence of the alpha-blocker phentolamine, 10(-5)M, and the beta-blocker propranolol, 10(-5)M, respectively; intracellular cyclic AMP levels and glycerol released into the buffer were measured. Consistent with past observations, epinephrine stimulated cyclic AMP and glycerol release when incubated with cells from fed subjects. Phentolamine enhanced this action and propranolol reduced it below the basal level. In contrast, epinephrine suppressed cyclic AMP and lipolysis when incubated with adipocytes from fasted individuals. This reversal in the effect of epinephrine on adipocy.se in alpha-activity. Additional studies suggest that this change in alpha- and beta-adrenergic receptor balance occurred after 1 day of fast and was not significantly exaggerated when fast was prolonged to 8 days. These findings are consistent with the view that in the fasting state the sympathetic nervous system and circulating catecholamines act to conserve adipose tissue triglyceride.


Hemodialysis hypotension is not the result of uremic peripheral autonomic neuropathy.

Five chronic hemodialysis patients with persistent hypotension during dialysis (MAP: 74.2 +/- 3.1 mm Hg) were given a number of standard tests of autonomic nervous system function and compared with eight normotensive hemodialysis patients (MAP: 96.4 +/- 3.4 mm Hg). Tests of efferent sympathetic nerves were normal in both groups, as were plasma catecholamine levels and the cold pressor test. The response to Valsalva maneuver and the venoconstriction reflex were generally abnormal and did not differentiate between the two groups. When adjusted for age and MAP, the baroreceptor slope to a high-pressure stimulus was diminished only in the hypotensive subjects. This result reinforces the previously described finding that many uremic patients do not develop a normal cardioacceleration during hypotension. Although reduced baroreceptor sensitivity may be a factor in the chronic hypotension of some hemodialysis patients, autonomic dysfunction alone is not a sufficient explanation of this phenomenon.


Chemotactic factor-induced superoxide radical generation by human neutrophils: requirement for proteinase (esterase) activity.

The requirement for proteinase (esterase) activity in the generation of O2- by human peripheral neutrophils was investigated. Neutrophils were activated by exposure to the chemotactic peptide FMLP and superoxide generation was assessed by ferricytochrome C reduction. Inhibition of O2- generation was observed by pretreating cells with the chloromethyl ketone derivatives of tosyl-phenylalanine (TPCK, ID50 2.2 X 10(-5)M) and tosyl-lysine (TLCK, ID50 1.9 X 10(-4)M). Dose-dependent inhibition was also noted with synthetic proteinase substrates, especially those of chymotrypsin-like specificity, phenylalanine and tryptophan methyl esters (ID50S approximately 1.5 X 10(-4)M), as compared to derivatives of basic amino acids, arginine and acetyl-lysine methyl esters, which caused negligible inhibition at 10(-3M. The inhibition of O2- generation demonstrated by TryME was time- and temperature-dependent and reversible with washing of the cells, whereas the inhibition seen with TPCK was irreversible. DFP at high concentrations, 10(-3)M or greater, caused moderate inhibition of O2- generation that was unaffected by exogenous serine and almost completely reversible by washing the cells. TPCK and TryME, at concentrations that caused marked inhibition of O2- generation, had no effect on the fmlp-induced calcium-45 uptake by the cells. These studies suggest that intact proteinase function is required for O2- generation and that this step follows the calcium influx in the activation sequence induced by FMLP.


Virology studies and Bell's palsy.

A retrospective analysis of 104 cases of facial nerve paralysis is presented, attention being focused on possible clues to link Bell's palsy with a virus infection. The findings are compared to those of another larger prospective series of cases. Some recent developments which suggest that Bell's palsy represents a reactivation of herpes simplex virus, rather than a primary infection, are discussed.


Traumatic aneurysm of the internal carotid artery and epistaxis. (Review of literature and report of a case).

A review of 108 years of world literature revealed 69 cases fulfilling the picture of a clinical syndrome of head injury, a latent period followed by epistaxis and cranial nerve lesions. A similar case has been added which is the fourth case to be reported to be due to a bullet injury. The applied anatomy of the intracranial internal carotid artery (ICA) and the aetiology, clinical syndrome, pathology, diagnosis, prognosis and management of ICA aneurysms have been briefly discussed.


The clinical evaluation of a new clobetasol propionate preparation in the treatment of otitis externa.

A new preparation containing clobetasol propionate, tetracycline and nystatin was compared with Otosporin in the the treatment of otitis externa. Thirty-two patients with bilateral disease and seventeen with unilateral disease completed the trial. Signs and symptoms were assessed by the clinician initially and at varying times after treatment. Both preparations were effective in relieving the symptoms with the majority of patients noticing improvement within seven days. Clobetasol propionate ear drops were significantly better than Otosporin when assessed on the patient's last visit to the hospital (p less than 0.05). There were less relapses in the clobetasol propionate group than in the Otosporin group.


Temporal oscillations in neuronal nets.

A model for the interactions of cortical neurons is derived and analyzed. It is shown that small amplitude spatially inhomogeneous standing oscillations can bifurcate from the rest state. In a periodic domain, traveling wave trains exist. Stability of these patterns is discussed in terms of biological parameters. Homoclinic and heteroclinic orbits are demonstrated for the space-clamped system.


Existence and stability of local excitations in homogeneous neural fields.

Dynamics of excitation patterns is studied in one-dimensional homogeneous lateral-inhibition type neural fields. The existence of a local excitation pattern solution as well as its waveform stability is proved by the use of the Schauder fixed-point theorem and a generalized version of the Perron-Frobenius theorem of positive matrices to the fuction space. The dynamcis of the field is in general multi-stable so that the field can keep short-term memory.


Adrenocorticotrophin-related peptides in adult and foetal sheep pituitary glands.

Differences in foetal and adult adrenal function may be due to qualitative as well as quantitative changes in the pituitary corticotrophic stimulus. Pituitary glands from adult and foetal sheep were freshly dissected and stored at -70 degrees C until extracted at pH 1.5. The extracts were subjected to chromatography on Sephadex G-100 superfine and fractions were assayed by multiple radioimmunoassays directed against the NH2- and CO2H-terminal sequences of ACTH and lipotrophin (LPH). Peaks corresponding to beta-melanocyte-stimulating hormone (beta-MSH), beta-LHP, gamma-LPH, beta-endorphin and ACTH were identified, with little or no evidence for the presence of alpha-MSH and corticotrophin-like intermediate lobe peptide. Three peaks of large molecular weight material, A. B and C, were identified and their relative proportions shown to be considerably greater in the foetus than in the adult. The immunoassay profile of peaks A and B suggested that they were 'stem hormones' which could give rise to a family of biologically active peptides. Since the 'family tree' which they engender varies according to the stage of development, it is proposed that the changes in the 'trophic family' may explain the different adrenal responses of the foetal and adult sheep.


Priming effect of luteinizing hormone releasing factor in vitro: role of protein synthesis, contractile elements, Ca2+ and cyclic AMP.

The mechanism of the priming effect of luteinizing hormone releasing factor (LH-RF) upon gonadotrophin secretion was studied using short-term incubation of hemipituitary glands from pro-oestrous rats. The dependence of the priming, but not the LH releasing action of LH-RF on protein synthesis in pituitary tissue was confirmed. Cytochalasin B failed to affect the first response to LH-RF, but abolished the priming effect, suggesting that the integrity of cellular microfilaments was essential. Colchicine and vinblastine did not modify the response to LH-RF. Neither inhibitors of DNA nor the inhibitor of RNA polymerase II, alpha-amanitin, significantly affected the priming action of LH-RF. Normal extracellular concentrations of Ca2+ were necessary for gonadotrophin release, but the priming effect was not significantly affected by low extracellular Ca2+ and could not be elicited by raising intracellular Ca2+ concentrations. Adenosine 3':5'-cyclic phosphate did not appear to act as a second messenger for either the gonadotrophin releasing or the priming action of LH-RF.


Do hypothalamic regulatory factors other than luteinizing hormone releasing factor exert a priming effect?

Extracts of hypothalamic tissue were applied to pituitary tissue in vitro to see whether they could exert a priming effect on gonadotrophin, adrenocorticotrophin, thyrotrophin or prolactin secretion. A clear-cut priming effect was seen only for gonadotrophin secretion.


An extra-adrenal action of adrenocorticotrophin: physiological induction of lipolysis by secretion of adrenocorticotrophin in obese rabbits.

The effect of minor stressful stimuli (saline infusions or venepunctures) on lipid mobilization (estimated by plasma free fatty acids (FFA) and glycerol levels) was studied in two strains of conscious rabbits of various ages. Infusions of 0.9% (w/v) NaCl solution induced a prompt increase of FFA levels in naturally obese rabbits while this effect was absent in younger lighter animals. Administration of propranolol (a beta-adrenoceptor blocking drug) beforehand did not prevent lipid mobilization. Adipose tissue in vitro was completely unresponsive to the lipid mobilizing action of adrenaline. The initial output of ACTH induced by stressful stimuli was quickly followed by the increase in the plasma concentration of FFA. A highly significant correlation (r = 0.84 or 0.85; P less than 0.01) existed between plasma ACTH and FFA levels 30 and 60 min after the start of stress. Dexamethasone completely prevented lipid mobilization while adipose tissue in vivo and in vitro responded strongly to ACTH. Consequently the lack of response of plasma FFA concentrations in corticosteroid-treated rabbits seemed to be linked to the failure of the pituitary gland to mobilize ACTH rather than to the unresponsiveness of adipose tissue to the pituitary hormone. Our data indicate that in the rabbit, under conditions of stress, ACTH is of greater physiological importance than catecholamines in the control of lipolysis.


Secretion of aldosterone in response to histamine in hypophysectomized-nephrectomized dogs.

In hypophysectomized-nephrectomized dogs after intravenous injection of histamine, a marked increase was observed in the rate of secretion of aldosterone, although it was smaller than that in intact dogs. Hypophysectomy plus bilateral nephrectomy greatly impaired the secretion of corticosterone and cortisol in the dog in response to histamine. However, a small yet significant increase in corticosterone and cortisol secretion was observed in hypophysectomized-nephrectomized dogs after intravenous injection of histamine. Additional experiments showed that plasma concentrations of potassium and sodium in hypophysectomized-nephrectomized dogs remained unchanged after intravenous injection of histamine. These results suggest that histamine stimulates aldosterone secretion in the dog partly by a direct effect on the adrenal cortical cells, whereas the effect of histamine on corticosterone and cortisol secretion is mediated mainly, but not totally, by pituitary release of ACTH.


Anaerobic rat heart: mitochondrial role in calcium uptake and contractility.

In order to evaluate the manner by which fumarate enhances contractility in the anaerobic heart, we examined Ca++ movements in isolated heart mitochondria and in the isolated perfused heart. Our experiments showed that in isolated antimycin A plus cyanide treated mitochondria: (a) Ca++ uptake was promoted by electron transport generated by fumarate-dependent oxidation of NADH, (b) Ca++ stimulated fumarate-dependent oxidation of NADH, (c) the ratio of Ca++ uptake:NADH oxidized was 1.7 and (d) the Ca++ sequestered is transiently highly mobile and is rapidly released upon collapse of the membrane potential. In anaerobic hearts perfused with glucose plus fumarate, malate and glutamate Ca++ levels were the same as those in oxygenated hearts while in anaerobic organs perfused with or without glucose Ca++ content was appreciably lower. Succinate production in anaerobic heart perfusions was related to: (a) an increased retention of Ca++ by the heart, (b) a diminution in peak aortic pressure generated by cardiac contractions and (c) an increase in heart rate. The information obtained indicates that mitochondria have a capability for Ca++ movement which be used physiologically, particularly in fumarate perfused anaerobic hearts, to assist the mechanism for contraction and relaxation of the heart.


Hormone-induced pigment translocations in amphibian dermal iridophores, in vitro: changes in cell shape.

Hormone-induced pigment translocation studies were conducted at both the light and electron microscopic levels on cultured dermal iridophores from the Mexican leaf frog, Pachymedusa dacnicolor. Two distinct types of dermal iridophores were characterized which differed in (1) their in vivo locations, (2) their overall morphologies in vitro, (3) their responses to alpha-MSH, ACTH, c-AMP or theophylline, (4) their physical alterations of light, and (5) certain ultrastructural features. One iridophore (Type I) was found to be physiologically responsive to the above hormones or agents by a reversible retraction of cellular processes and a thickening of the cell body, an event which is inhibited by cytochalasin B. The other iridophore (Type II) appeared to be unresponsive. Type I iridophores contain cube-like pigmentary organelles, refractosomes, while Type II iridophores contain larger, bar-shaped refractosomes. In addition, both iridophore types contain 60  and 100 A microfilaments as well as microtubules. By in large, micorfilaments were found within microvilli, beneath and parallel to the plasma membrane and in the perinuclear region. Occasionally, bundles of 100 A microfilaments were found between layers of refractosomes in Type I iridophores. These results are discussed in relation to hormone-induced changes in cell shape.


The effect of collagen on the cyclic AMP content of embryonic somites.

Previous studies have demonstrated that collagen substrates stimulate in vitro somite chondrogenesis, and that agents that elevate intracellular cyclic AMP levels in hibit the ability of somites to respond to the inductive influence of collagen. In the present investigation, radiommunoassay was utilized to compare the cyclic AMP content of somite explants cultured on purified Type I collagen substrates with control explants cultured on Millipore filters. During the period of culture, the cyclic AMP content of collagen-treated explants is significantly lower than the cyclic AMP content of control explants. The cyclic AMP content of collagen-treated explants is 66% of control values as early as one hour following the initiation of culture, and the cyclic AMP content of collagen-treated explants remains lower than controls throughout the 3-day cultured period. The greatest difference in the cyclic AMP content of collagen-treated and control explants is observed at the seventeenth hour of culture, at which time the cyclic AMP content of collagen-treated explants is 56% of controls. These results combined with previous studies provides support for the hypothesis that collagen elicits a reduction in the cyclic AMP content of embroyic somites and that this reduction is necessary to trigger chondrogenic differentiation.


Dopamine blockade of the thyroliberin -- induced cyclic AMP accumulation in rat anterior pituitary.

Effects of dopamine and thyroliberin on the cyclic AMP concentration in the rat anterior pituitary gland were investigated in vitro. Dopamine by itself had no effect on the anterior pituitary cAMP concentration but inhibited the cAMP-accumulating effect of thyroliberin.


Qualitative aspects on motility changes in mice induced by low doses of apomorphine and clonidine.

Single mice were treated with apomorphine (0.05, 0.2 or 0.8 mg/kg) or clonidine (0.025 or 0.05 mg/kg), and were placed in a new kind of motility meter, allowing classification of movements in terms of size, and also charting the motility with the help of an X/Y pen recorder. All treatments induced reduction of motility in this model. Apomorphine, 0.8 mg/kg, considerably changed the motility type, whereas the lower doses did not. This finding is consistent with earlier studies on gross lobomotor activity and suggests that the higher dose alone evokes a significant direct net stimulation of postsynaptic dopamine receptors, whereas the lower doses merely reduce the physiological stimulation of these receptors by inhibiting dopamine neurotransmission. Clonidine, 0.025 mg/kg, depressed motility at least as much as did 0.05 mg/kg, in agreement with previous behavioural studies.


The effects of the combined administration of gamma-hydroxybutyrate and diazepam on sleep parameters in the rat.

The actions of diazepam and gamma-hydroxybutyrate (GHB) were assessed on the sleep-wakefulness cycle of male Wistar rats. One and 2 mg/kg diazepam and 12.5 mg/kg GHB had no effects on the sleep variables. After 25 mg/kg GHB, slow wave sleep (SWS2) was significantly increased. Following the combined administration of non-effective doses of GHB and diazepam, significantly higher amounts of SWS2 at the expense of wakefulness were obtained. The injection of a subconvulsant dose of bicuculline (2.5 mg/kg) prior to treatments which significantly increased SWS2 prevented this effect to show up. It is suggested that the actions of GHB and diazepam on the sleep-awake cycle are related to the same neurotransmitter system.


Arginine vasotocin as a pineal hormone.

The pineal nonapeptide hormone arginine vasotocin (AVT) is synthesized by the ependymal cells of the pineal recess and subcommissural organ and stored in so far undefined cells of the pineal gland proper. AVT is first released into the cerebrospinal fluid (CSF) and reaches the blood only secondarily after its absorption from CSF. It displays a diurnal rhythm in the pineal and CSF, suggesting its release into the CSF during the night in the dark. Melatonin represents its releasing hormone. AVT exerts both its endocrine and non-endocrine effects by a unique mechanism involving the activation of serotonin neurotransmission in the brain with resultant inhibition of release of hypothalamic releasing and inhibiting hormones and induction of sleep. It produces both its endocrine effects and sleep at concentrations equivalent to only several hundreds of molecules, being thus by far the most active hormone so far known. Midbrain raphe nuclei or some structures intimately correlated with these cell bodies, most contain the extremely sensitive and specific AVT receptors in the mammalian brain. In contrast with its natural analogues arginine vasopressin and oxytocin which are mainly blood hormones, AVT is a CSF hormone whose major if not the sole site of action is the brain itself.


Sensitivity and cyclic nucleotides in the rat pineal gland.

beta-Adrenergic stimulation induces serotonin N-acetyltransferase (SNAT) activity in the rat pineal gland. The magnitude and some of the characteristics of this response vary as a function of the gland's previous exposure to stimulation. A period of stimulation results in a subsensitive response to subsequent stimulation. A period without stimulation provides a supersensitive response to subsequent stimulation. Investigations concerned with the mechanisms regulating the rat pineal's sensitivity to beta-adrenergic stimulation are described. These have focused on the regulation of cyclic AMP metabolism. Several of the components involved in the induction of SNAT activity appear to participate in the regulation of sensitivity. These include the beta-adrenergic binding sites, the catecholamine-sensitive adenylate cyclase, the cyclic nucleotide phosphodiesterase, and the cyclic AMP-dependent protein kinase. Thus, the rat pineal's sensitivity to beta-adrenergic stimulation appears to be regulated at multiple sites. Other investigations have focused on the regulation of pineal cyclic GMP metabolism. Unlike cyclic AMP, the stimulation of cyclic GMP synthesis requires the presence of intact nerve endings and of extracellular calcium. Some of the characteristics of pineal cyclic GMP regulation are described.


Developmental plasticity of central noradrenaline neurons after neonatal damage--changes in transmitter functions.

The effects of neonatal 6-hydroxydopamine (6-OH-DA) treatment (systemic administration) on noradrenaline (NA) metabolism, turn over, and receptor characteristics have been investigated in rat brain in the adult stage. This treatment is known to preferentially affect the locus coeruleus (LC) NA system leading to a marked NA denervation in the central cortex and hyperinnervation of NA nerve terminals in the pons and medulla oblongata without influencing the LC perikarya. The main NA metabolite, 3-methoxy-4-hydroxy-phenylglycol (MOPEG) was reduced by about 70% in the cerebral cortex after 6-OH-DA treatment at birth while the endogenous NA was almost completely depleted (-92%). The MOPEG levels were not significantly changed in the pons medulla after 6-OH-DA treatment in contrast to the 60% increase of the endogenous NA concentration. The relative reduction of NA in the cerebral cortex of 6-OH-DA treated rats increased in the cerebral cortex following administration of the tyrosine hydroxylase inhibitor alpha-methyl-p-tyrosine (H44/68) compared to the control, while the H44/68 induced depletion of NA was reduced in the pons medulla after 6-OH-DA. The steady-state level of endogenous NA and the effect of H44/68 were unchanged in the LC perikarya after 6-OH-DA treatment. These results indicate that the NA turn over in remaining NA nerve terminals in the cerebral cortex is increased after 6-OH-DA, while decreased in the pons-medulla, possible related to changes in the activation of presynaptic alpha-adrenoreceptors in both regions. NA-induced formation of cAMP in vitro was found to be markedly increased in the cerebral cortex after 6-OH-DA, whereas no consistent change was observed in the pons medulla. Measurements of alpha- and beta-receptor binding in vitro using radioligand techniques showed an increase of binding sites (20%--50%) for both receptors in the neocortex aster 6-OH-DA, whereas no changes were observed in the pons medulla. The 6-OH-DA induced changes in NA turnover, cAMP generating systems, and receptor density may all represent compensatory processes following the altered development of the NA neurons induced by 6-OH-DA.


Peripheral neuropathy in angioimmunoblastic lymphadenopathy with dysproteinaemia.

A case of angioimmunoblastic lymphadenopathy with dysproteinaemia complicated by a subacute peripheral neuropathy is described. Clinically  the neuropathy was mainly motor, but pain and paraesthesiae in the legs were also present. Sural nerve biopsy indicated decreased numbers of myelinated fibres, mainly the largest, without actual signs of degeneration or regeneration. However, the occurrence of denervation bands indicated that degeneration had led to loss of myelinated fibres.


Ataxia-telangiectasia. Report of a case with Lewy bodies and vascular abnormalities within cerebral tissue.

The neuropathologic findings in a 31 year old patient with ataxia-telangiectasia are presented. There was an unusually severe and widespread distribution of the recognized abnormalities within the nervous system. In addition, there were Lewy bodies in the substantia nigra and widely distributed vascular changes in cerebral tissue. The nature of the vascular changes is discussed. These findings underline and expand the known spectrum of nervous system involvement in this condition.


Ultrastructural study of zinc pyridinethione-induced peripheral neuropathy.

This study describes, for the first time, the neuropathy of zinc pyridinethione (ZPT) toxicity in rats. Although hind-limb weakness has been previously reported as a consequence of dietary ZPT ingestion in rats and rabbits, the cause of the paralysis has been unexplained. Sequential morphologic studies in rats fed a diet containing 166 ppm of ZPT revealed a dying-back neuropathy characterized by the early accumulation of tubulo-vesicular profiles in the motor nerve terminals. Continued exposure resulted in similar abnormalities in the intrasmuscular nerves and later in the peroneal and posterior tibial nerves and nerve branches to individual muscles. Axonal degeneration and regeneration followed the initial pathologic changes. There was relative sparing of the sensory nerve terminals of the muscle spindles. Central nervous system axons in the long descending tracts of the spinal cord and in the cerebellar vermis showed similar but quantitatively fewer axonal changes compared to the peripheral nerves. The central effects occurred only after prolonged administration of ZPT.


Cytologic observations on axotomized feline Betz cells. II. Quantitative ultrastructural findings.

Quantitative electron microscopic examination was made of Betz cells of two unoperated cats as well as cats subjected to left lateral funiculotomy 5, 10, 28 and 49 days before sacrifice. The percent cytoplasmic composition of chromatolyzed, right-sided Betz cells contributed by cisternal elements of RER, Golgi apparatus and dense bodies and the percent perikaryal membrane apposed by subsurface cisterns were unchanged from the normal despite marked qualitative alterations of the cytoplasm. However, 49 days postoperatively mitochondrial numerical density of axotomized, right-sided Betz cells was significantly less than at 0, 10 and 28 days post funiculotomy. Importantly, normal-appearing Betz cells ipsilateral to corticospinal tract section showed an increase in mitochondrial numerical density 5 days postoperatively. Operation did not induce change in the % perikaryal coverage by axosomatic boutons. Retraction of axosomatic boutons, though often reported for other neuronal populations undergoing axon reaction, is not a necessary feature of the axon reaction of feline Betz cells.


Experimental interfascicular nerve grafting.

Twenty-nine adult rhesus monkeys underwent complete laceration of both tibial nerves at mid-thigh level and repair by different methods to study the relative efficacy of autogenous interfascicular nerve grafts. Sixteen animals in Group I had an interfascicular graft repair using short sural nerve autografts on one limb and fascicular repair without grafts on the other limb. Thirteen animals in Group II, after having a 1-cm segment of tibial nerve resected, had an interfascicular graft repair without tension in one limb and an epineurial repair under moderate tension in the other limb. Evoked nerve and muscle action potentials and muscle strenght in response to repetitive and tetanic stimulation were recorded as baseline values prelaceration and then on re-exploration at 4, 6, 9, or 12 months. All nerves were examined by light and electron microscopy. Electrophysiological data, particularly muscle strength response, showed non-graft repairs to be superior at 4 and 6 months of regeneration. However, by 9 and 12 months the graft repairs had caught up and were equal to the non-graft repairs. Histologically, it was observed that many axons missed the graft segments and were present in extrafascicular connective tissues. Nonetheless, enough axons regenerated to the distal nerve to explain the success of these relatively short grafts. From the results of these experiments, it is concluded that use of autogenous interfascicular grafts offers no advantage over end-to-end non-graft repair. When and end-to-end repair cannot be achieved, use of short interfascicular nerve grafts is feasible and will work.


Histological studies of experimental interfascicular graft repairs.

Biopsies of sutured and grafted primate peripheral nerves were examined by light and electron microscopy after the final set of electrical measurements had been recorded. Inspection of all proximal stumps showed the expected regenerative activity which was not affected by the nature of the nerve repair. Transverse sections through the epineurial, interfascicular, and graft suture lines showed a similar pattern in all animals and at this site nerves sutured by epineurial technique could only be distinguished from those sutured by fascicular technique by loci of the non-absorbable suture. Fascicular repairs, whether done fascicle-to-fascicle or with interposition of grafts, had a more lengthy neuroma than did the epineurial repairs. Maintenance of fascicular architecture through the course of the grafts was variable. Fascicular structure was frequently absent in the central graft segments and in segments close to the second suture site. The method of repair used more proximally could not be distinguished by evaluation of distal stump segments. Measurements of myelinated fiber size made of distal stump axons revealed no statistical difference between the methods of repair.


Atypical fibrous histiocytoma of the thoracic spine. Case report.

A 14-year-old boy presented with mid-thoracic pain, leg weakness, and bladder dysfunction. Plain x-ray films and bone scan were normal whereas myelography demonstrated an extradural mass extending from T-2 to T-5. Exploration revealed a large tumor adherent to dura and bone. Pathological verification of an atypical fibrous histiocytoma prompted the report of this rare tumor of the vertebral column.


Longitudinal study of tolyl-reactive IgE antibodies in workers hypersensitive to TDI.

Three workers with TDI hypersensitivity were evaluated for IgE antibodies to TDI over a period of 13 months. A radioallergosorbent test (RAST) system was employed using p-toly(mono)isocyanate-human serum albumin antigen covalently bound to cyanogen bromide-activated paper discs. IgE antibody titers were consistently elevated in two individuals who experienced several bronchial hypersensitivity responses to TDI during the study period. The responses were either solely asthmatic or asthmatic accompanied by cutaneous hypersensitivity reactions. By contrast, antibody titers in a third subject who had not experienced any hyersensitivity reactions during the study period continually decreased, falling to insignificant levels after 12 months. In the absence of renewed TDI exposure, sensitive workers may have titers indistinguishable from those of workers exposed to TDI but without sensitivity to the chemical.


Effects of chronic amorphous silica exposure on sequential pulmonary function.

To assess clincial effects of precipitated amorphous silica (PAS), the authors reviewed serial spirograms, respiratory questionnaires, and chest radiographs of 165 workers exposed for a mean of 8.6 years. Monthly exposure was graded on a 1 to 4 scale and a "cumulative exposure index" (CEI) calculated for each worker from the sum of measured exposure. A "mean exposure index" (MEI) was calculated by dividing the CEI by total months exposed. Sputum production and dyspnea were inversely correlated with CEI, while cough and dyspnea correlated with mean pack-years of smoking but not PAS exposure. Linear regression analysis of yearly change of all pulmonary function variables (FVC, FEV1, FEV1/FVC, FEF25-75) showed no correlation with either the dose of PAS (CEI) or total years of exposure. Among 44 workers with a mean exposure time of 18 years (range 10-35 years), yearly decline of FVC and FEV1 were similar to the overall group. Of 143 workers with serial radiographs and exposure to only PAS, none had radiographic pneumoconiosis. Respiratory symptoms in PAS workers correlate with smoking but not with PAS exposure, while serial pulmonary function values and chest radiographs are not adversely affected by long-term exposure.


Cadmium nephropathy--a clinical evaluation.

Decreased GFR and TmG and increased beta-2-microglobulin excretion were found in a worker with a nine-year exposure to cadmium. The patient's recent marked weight loss and other multiple symptoms could not be explained on these abnormalities or by results of other specialized studies. Increased urinary beta-2-microglobulin without proteinuria but with elevated blood and urine cadmium levels in this patient raises questions about which biological indicators will prove to be most helpful in effectively monitoring cadmium workers.


Extrapulmonary manifestations of adenovirus type 7 pneumonia simulating Reye syndrome and the possible role of an adenovirus toxin.

Three children developed extensive extrapulmonary disease in the course of fatal adenovirus type 7 pneumonia. Several clinical features, including the unexpected onset of coma, suggested the development of Reye syndrome, but biochemical and histopathologic findings were inconsistent with this diagnosis. Virologic and pathologic studies did not reveal evidence of extrapulmonary adenovirus infection, despite clinical involvement of the liver, skeletal muscle, and central nervous system. The detection in premortem sera from all three patients of adenovirus penton antigen, known to be cytotoxic in vitro, suggests a possible mechanism for the production of extrapulmonary pathology in the absence of extrapulmonary virus infection.


Cytomegaloviruria in older infants in intensive care nurseries.

Over a four-month period, urine specimens for viral isolation were obtained weekly from all infants older than three weeks in two intensive care nurseries. These babies comprised 43% of the patients in the nurseries surveyed. Cytomegalovirus was cultured from 13 of 93 (14%) of these infants. Eleven of 13 infants who developed cytomegaloviruria were born prematurely, and nine of these 11 were found to be excreting CMV before they reached 40 weeks postconception. Infants excreting CMV received blood transfusions from a mean of 10.45 (+/- 1.80 SE) different donors versus 5.10 (+/- 0.55 SE) for infants without viruria (P less than 0.002) and five of 14 infants undergoing one or more exchange transfusions developed cytomegaloviruria (P less than 0.05). The possible role of other CMV reservoirs and the importance of these findings are discussed.


The actions of excitatory amino acids on motoneurones in the feline spinal cord.

1. Combined recording or ionophoretic electrodes of the concentric type were used to investigate the depolarizing responses of DL-homocysteate (DLH) and L-glutamate in cat lumbar motoneurones. 2. Typically, DLH responses were slow both in onset and recovery, while glutamate responses were fast in onset and recovery and were frequently accompanied by a post-response hyperpolarization. 3. DLH responses (smaller than those necessary to evoke firing) were accompanied by a stable decrease in GM. This decrease was usually more than could be accounted for by anomalous rectification of the membrane. 4. Small glutamate responses were accompanied by either a small decrease, no change or a small increase in GM. There was a biphasic change in GM during large responses: GM decreased during the rising phase and early part of the response plateau and thereafter increased as the depolarization was maintained. It is proposed that the high conductance state during glutamate application (but not the depolarization itself) is a manifestation of glutamate uptake. 5. Firing evoked by DLH was stable during very long applications of the drug. Firing evoked by glutamate was usually of short duration, despite the maintained depolarization. 6. No reversal potential for the DLH responses could be demonstrated, but the responses decreased in size both with hyperpolarization and depolarization of the membrane. A 'null point' of the response in the negative direction was found to be approximately -95 mV. 7. DLH resonses were insensitive to changes in the internal Cl concentration. When the external K concentration was increased by K+ ionophoresis, the DLH responses became smaller. It is concluded that the DLH response is probably mediated via a decrease in K+ conductance and that the availability of this conductance channel is potential dependent. 8. Changes in the sizes of evoked potentials (e.p.s.p.s, i.p.s.p.s and a.h.p.s) with DLH and glutamate responses were investigated. The size of each of these evoked potentials was inversely related to GM during the responses; thus they all showed stable increases during DLH responses. E.p.s.p.s recorded during DLH were of longer half-width and time-to-peak than the control, but there was no change in the maximum slope (V.sec-1). When e.p.s.p.s decreased in size with glutamate the time-to-peak remained constant. 9. Acidic amino acids have been implicated as natural excitatory transmitters. The consequence of our results for the mechanism of excitatory transmission is therefore discussed.


Transmission and contraction fatigue of rat motor units in relation to succinate dehydrogenase activity of motor unit fibres.

1. The fatigue in rat anterior tibial (a.t.) motor units was studied and related to microphotometric determinations of succinate dehydrogenase (SDH) activity of the motor unit muscle fibres. 2. Anterior tibial contains fast-twitch type II fibre units with an average contraction time of 11 msec and about 5% slow-twitch type I fibre units with an average contraction time of 20 msec. 3. In type II fibres stained for SDH, absorbance varied continuously from 0.046 to 0.569 and inversely to fibre size, except for the largest fibres. 4. Resistance to fatigue of fast motor units to 100 Hz intermittent stimulation varied continuously within a wide range in near linear relations to absorbance for SDH of unit fibres and inversely to tetanic tension, except for motor units with the largest fibres and the largest tetanic tension. 5. Neither resistance to fatigue nor SDH activity lent itself to any categorization of motor units or fibres into well demarcated functional or histochemical types, since both parameters varied continuously in the unit and fibre population of the muscle. 6. The direct relation between resistance to fatigue of fast-twitch motor units and SDH activity of unit fibres appeared valid for fatigue resistance of: (a) neuromuscular transmission, tested with 100 Hz intermittent stimulation which gave concomitant failure of electrical and mechanical response, (b) excitation--contraction coupling,  demonstrated by post-stimulatory depression of twitch tension with preserved maximum tetanus tension and action potential, and (c) contractile mechanism; excitation--contraction coupling?, tested with low frequency stimulation which gave decline of twitch and maximum tetanus tension with preserved action potential. 7. It is suggested that the endurance of each link in the chain of events leading to contraction, including neuromuscular junction and the excitation--contraction coupling system, is under aerobic conditions matched to the contractile capacity of the fibre expressed by its oxidative enzyme activity.


Some effects of hypertonic solution on the properties of spontaneous transmitter release in the hypogastric ganglia of guinea-pigs.

1. The effects of hypertonic solutions on spontaneous transmitter release at synapses in the hypogastric ganglia of guinea-pigs were studied using intracellular recording techniques. 2. Hypertonic solutions were found to cause a marked hyperpolarization of the ganglion cells and this was accompanied by an increase in both the threshold and input resistance of these cells. 3. The frequency of spontaneous quantal release was found to depend exponentially on the tonicity of the bathing solution with a 50% increase in tonicity producing a 7-9 fold increase in the frequency of release. 4. Hypertonic solutions were also found to increase the degree of bursting within the spontaneous process so that the proportion of multiquantal spontaneous potentials was found to be increased in such solutions. 5. Two different models, the 'charge-screening' model (Van der Kloot & Kita, 1973) and the 'Ca-increase' model (Shimoni, Alnaes & Rahamimoff, 1977), for the mechanism underlying the effects of hypertonic solution were considered.


Dorsal root potentials and changes in extracellular potassium in the spinal cord of the frog.

1. In the present study changes in extracellular potassium, ([K]e), were recorded in the isolated spinal cord of the frog with glial cell recordings and K-selective micro-electrodes to test the hypothesis that elevations in [K]e during neuronal activity produce the dorsal root potential (d.r.p.). 2. Sucrose gap recording from the dorsal root (d.r.) was used to record responses to root stimulation and to exogenously applied K+. 3. Stimulation of the ventral root, which elicits a d.r.p. in the frog spinal cord, was not associated with any change in [K]e, suggesting that d.r.p.s produced by ventral root stimulation are not due to changes in [K]e. 4. The largest change in [K]e observed following single stimuli to the dorsal root was 0.4 mM. Such a change in [K]e, if evenly distributed, would depolarize the dorsal root by about 1 mV and yet the simultaneously recorded d.r.p. evoked by stimulating an adjacent dorsal root (d.r.-d.r.p.) was over 10 mV. 5. The time-to-peak of the glidal cell responses was 10 times that of the d.r.-d.r.p. Low frequency (1-10 Hz) d.r. stimulation caused a decremental summation of glial cell responses, while there was no summation in the d.r.-d.r.p. These results suggest that the d.r.p. produced by single d.r. stimulation is generated in large part by a mechanism other than a change in [K]e. 6. During high frequency d.r. stimulation, which evoked 6-8 mM increases in [K]e, the adjacent d.r. was depolarized to a greater extent than that produced by single stimuli. The magnitude of this depolarization was similar to that produced by applying a [K]e equivalent to that observed in the spinal cord during high frequency stimulation. Thus, a substantial component of the sustained d.r. depolarization during high frequency d.r. stimulation may result from changes in [K]e. 7. In the presence of magnesium, high frequency d.r. stimulation evoked a picrotoxin resistant depolarization of an adjacent d.r. whose magnitude correlated well with the changes in [K]e recorded in the spinal cord. 8. In the presence of picrotoxin a slow, long duration depolarization of the d.r. occurred following single stimuli to the adjacent d.r. and the appearance and time course of this response correlated well with the time course of changes in [K]e. 9. Addition of K+ to the Ringer solution in concentrations up to 12 mM had a facilitatory action on reflex activity in the frog spinal cord. 10 The present results suggest that although changes in [K]e play a relatively minor role in generating d.r.p.s. elicited by single d.r. stimulation, the sustained dorsal root depolarization evoked either by high frequency stimulation or by single stimuli in the presence of picrotoxin may be due to a considerable extent to [K]e.


Relation between catecholamine-induced cyclic AMP changes and hyperpolarization in isolated rat sympathetic ganglia.

1. The effect of catecholamines on cyclic adenosine 3'5'-monophosphate (cyclic AMP) production in isolated rat superior cervical ganglia has been measured under experimental conditions in which they also produce ganglion hyperpolarization.2. (+/-)Isoprenaline (1 muM) increased cyclic AMP levels by 8-100 times after 15 min incubation at 25 degrees C. Half-maximal stimulation occurred at about 0.03 muM. This was due to stimulation of beta-receptors, since it was prevented by 1 muM-propranolol but not by 1 muM-phentolamine.3. The alpha-agonists phenylephrine (100 muM), dopamine (100 muM) and clonidine (1 muM) did not produce a detectable increase in ganglionic cyclic AMP. Dopamine (100 muM) was also ineffective at 37 degrees C in the presence of 10 mM-theophylline.4. Exogenous cyclic AMP (0.01-1 mM) hyperpolarized the ganglion. This effect was replicated by other adenosine compounds, most effectively by adenosine and by adenosine 5'-monophosphate, and was antagonized by theophylline. Dibutyryl cyclic AMP was weaker than cyclic AMP.5. Neither theophylline nor the non-xanthine phosphodiesterase inhibitor, Ro 20-1724, enhanced the hyperpolarizing actions of noradrenaline or dopamine.6. Since catecholamine-induced hyperpolarization of the isolated rat ganglion is induced via alpha-receptors, whereas cyclic AMP-production is induced via beta-receptors, it is concluded that cyclic AMP is unlikely to mediate the hyperpolarization. The effect of exogenous cyclic AMP may be due to an action on external adenosine-receptors.


Voltage clamping of unparalysed cut rat diaphragm for study of transmitter release.

1. As a result of the cutting procedure there is a decrease in the membrane potential, input resistance, and space constant of the rat diaphragm, and there is no contraction when the phrenic nerve is stimulated. While the reversal potential of  end-plate currents in cut preparations appears normal, the size of miniature end-plate currents (m.e.p.c.s.) is slightly decreased. 2. An increase in the external concentration of potassium from 2.5 to 10.0 mM results only in a minor change (less than 5%) in statistical parameters of transmitter release (m, n and p). The size of m.e.p.c. is also almost unchanged. 3. A decrease in temperature from 37 degrees to 15 degrees C resulted in a decrease of m, n and p; however, the values are less than 25% smaller at 25 degrees C than at 37 degrees C. The size of m.e.p.c.s is very insensitive to changes in temperature. 4. As previously reported for the frog neuromuscular junction, changes in muscle membrane potential of cut and uncut rat diaphragm due to voltage clamping affect the frequency of m.e.p.c.s only in medium with raised external K concentration. The dependence of frequency of m.e.p.c.s on muscle membrane potential is remarkably similar in cut and uncut preparations. Evoked release in cut preparation in normal medium is not affected by the change in muscle membrane potential.


Change of statistical parameters of transmitter release during various kinetic tests in unparalysed voltage-clamped rat diaphragm.

1. The statistical nature of transmitter release was studied in unparalysed cut rat diaphragm using a voltage clamp technique at room temperature (23 degrees C). 2. While the binomial distribution described observed amplitude histograms well, the Poisson distribution was clearly inadequate. Values of m ranged from 40 to 45, while values for n varied from 45 to 53 and p was between 0.82 and 0.90. 3. Estimation of the probability of release from the transient decay of e.p.c.s in short tetanic trains (11 pulses, 150 Hz) gave values of p from 0.031 to 0.054, which are more than one order of magnitude lower than statistical estimates. 4. As a result of the short tetanic stimulation (5 pulses, 20, 50 and 100 Hz) there is an initial transient facilitation which afterwards becomes masked by depression. Statistical analysis suggests that the changes in the average numbers of quanta released (m) could be attributed to the change in the immediately available store (n). 5. During long tetanic stimulation (4000 pulses, 10-100 Hz) statistical analysis suggests that the decrease in the average number of quanta released (m) could be attributed almost entirely to the decrease in the immediately available store (n). The probability of release (p) decreased only slightly. The extent of the post-tetanic potentiation indicates that it cannot be explained on the grounds of increased probability of release (p) only. There should be an increase as well in the immediately available store (n). 6. It is suggested that while depression is most likely caused by the depletion of the immediately available store due to insufficient replenishment, the facilitation is probably caused by the increase in the capacity of the immediately available store to contain transmitter.


Presynaptic action of curare.

1. As a result of a conditioning phrenic nerve stimulus, end-plate currents (e.p.c.s) in a voltage clamped uncurarized cut diaphragm show a facilitation which reaches its maximum at 30-40 msec and subsequently decays with a time constant from 150 to 200 msec. In curarized (cut or uncut) diaphragms, however, the conditioning stimulus causes a depression which reaches its maximal value at 10 msec and then decays slowly with a time constant of about 3 sec. This indicates that curare strongly interferes with the process of transmitter release. 2. The presynaptic action of curare is also evident if short tetanic trains are given. In uncurarized preparations e.p.c.s decay in size much more slowly than in curarized preparations, and usually show a transient facilitation. 3. These results can be explained in terms of a model where curare blocks presynaptic depolarizing action of ACh. As a result of this presumed curare action a small increase in Ca permeability and subsequent entry of Ca associated with depolarization are also blocked, and the facilitation resulting from that entry of Ca is abolished.


Effects of cold adaptation and re-adaptation upon the mitochondrial phospholipids of brown adipose tissue.

1. A study of the mitochondrial phospholipids, phospholipid fatty acid patterns and enzyme activities was investigated in brown tissue (B.A.T.) from rats chronically exposed to cold and/or treated with thyroxine. 2. The total activities of the oxidative enzymes were increased after cold exposure, but not after thyroxine treatment. 3. Cold exposure increased the amount of phosphatidylethanolamine, phosphatidylcholine, cardiolipin and lysophospholipids, the effect being greatest for phosphatidylethanolamine. At the same time, there were marked alterations in the fatty acid composition of the mitochondrial phospholipids (decrease of palmitic, palmitoleic and oleic acids ; increase of stearic, linoleic and arachidonic acids). 4. All these cold-induced alterations were reversed by re-adaptation of the animal to a normal temperature range. 5. The alterations of the fatty acid composition of phospholipids could be explained by changes in the rate of individual fatty acid biosynthesis.


[Hypothalamic influence on differentiation of the immunoreactive ACTH beta-LPH, alpha- and beta-endorphin containing cells in adenohypophysial primordia of rat fetus in organ culture (author's transl)].

Adenohypophysial primordia were isolated in rat fetuses from day 12.5 to day 15.5 of gestation. The organ culture employed for maintenance of the primordia was made up according to Watanabe et al. (1973). The fixation of primordia in Bouin Hollande's solution was performed after 9, 8, 7 or 6 days of culture when the normal duration of pregnancy was achieved. The cultivated primordia were immunologically studied using different antisera: anti-alpha(17-39)ACTH, anti-beta(1-24)ACTH, anti-beta-LPH, anti-alpha and anti-beta-endorphins, with immunoperoxidase or immunofluorescence techniques, including control experiments of the specificity of the antisera. A similar study was performed on pituitaries removed from normal rat fetuses from day 16.5 of gestation and each day up to birth, and fixated immediately. In vivo the first cells reacting with all the antisera used in this study were observed on day 16.5 of gestation; their number increased during gestation (Fig. 1 A, B and C). Immunoreactive cells with the different antisera could be detected in primordia isolated on day 12.5 of gestation after 9 days of culture. Numerous groups of cells were observed in primordia of older fetuses (Fig. 2 A and B). These data indicate that the corticotropic cells in rat fetuses could start to be differentiated without stimuli from the hypothalamus since primordia were isolated before the appearance of this cell type in normal rat fetuses and before the differentiation of the hypothalamus. The presence of ACTH and other peptides such as beta-LPH or beta-endorphin would support the hypothesis of a common precursor in this cell type existing early in gestation. Similar results were obtained in human fetuses.