[Examination of plasma membrane-enriched fraction from guinea pig intestinal smooth muscle by means of some marker enzymes (author's transl)]. By means of a preparation technique based on the discontinuous sucrose density gradient, subcellular fractions were isolated from guinea pig intestinal smooth muscle cells. A fraction which distributed to a 33% sucrose layer showed relatively high activities of 5'-nucleotidase, Na+ . K+-ATPase and ouabain sensitive Na+ . K+-ATPase. The fraction had a low NaN3 sensitive Mg2+-ATPase activity. On the other hand, the high activity of glucose-6-phosphatase showed a broad distribution. Though the sucrose density gradient proceeded over a series of the fine layers, cross-contamination of microsome into the 33% sucrose fraction was not reduced. To reduce microsomal cross-contamination, another procedure was employed. The homogenization time of 77000 xg sediment to be layered on the top of the sucrose density gradients was prolonged. This procedure did not change the distribution of K+ activated p-nitrophenylphosphatase, K+ activated ouabain sensitive p-nitrophenylphosphatase and ouabain sensitive Na+ . K+-ATPase activities. The peak of NADH cytochrome c reductase activity was shifted to a 38% sucrose fraction from a 33% sucrose fraction and the activity of this marker enzyme in the 33% sucrose fraction decreased to 60% of that of the prior procedure.


Structure and function of H+-ATPase.

Structure and function of H+-ATPase. (1) Extensive studies on proton-translocating ATPase (H+-ATPase) revealed that H+-ATPase is an energy transforming device universally distributed in membranes of almost all kinds of cells. (2) Crystallization of the catalytic portion (F1) of H+-ATPase showed that F1 is a hexagonal molecule with a central hole. The diameter of F1 is about 90 A and its molecular weight is about 380,000. (3) Use of thermophilic F1 permits the complete reconstitution of F1 from its five subunits (alpha, beta, gamma, delta, epsilon) and demonstration of the gate function of the gamma delta epsilon-complex, the catalytic function of beta (supported by alpha and gamma), and the H+-translocating functions of all five subunits. (4) Studies using purified thermostable F0 showed that F0 is an H+-channel portion of H+-ATPase. The direct measurement of H+-flux through F0, sequencing of DCCD-binding protein, and isolation of F1-binding protein are described. (5) The subunit stoichiometry of F1 may be alpha 3 beta 3 gamma delta epsilon. (6) Reconstitution of stable H+-ATPase-liposomes revealed that ATP is directly synthesized by the flow of H+ driven by an electrochemical potential gradient and that H+ is translocated by ATP hydrolysis. This rules out functions for all the hypothetical components that do not belong to H+-ATPase in H+-driven ATP synthesis. The roles of conformation change and other phenomena in ATP synthesis are also discussed.


Mitochondrial function in cerebral ischemia and hypoxia: comparison of inhibitory and adaptive responses. In cerebral ischemia, brain oxygen supply is totally exhausted within seconds. This necessitates cessation of mitochondrial electron transfer and energy (ATP) production. After certain periods of ATP deficiency of from 5 to 90 min, irreversible damage of mitochondrial membranes occurs. This results in decreased mitochondrial function, characterized by inhibited State 3 respiratory rates, low respiratory control ratios, and inhibited Ca2+ transport activities. A 30-min recirculation period of the ischemic brain tissue induces total restitution of mitochondrial respiratory capacity after complete ischemia, but not after incomplete ischemia. Regional in situ measurements of brain pyridine nucleotide redox levels, tissue ATP, and lactate concentrations indicate variable metabolic responses of different brain regions to oligemia. Macroheterogeneity from region to region, as well as microheterogeneity within a region are demonstrated. Contrary to the effect of tissue ischemia involving reduced or zero cerebral blood flow and tissue oxygenation, sublethal hypoxia alone at normal or increased levels of blood flow induces adaptation of the mitochondrial enzyme system to a new level of respiratory capacity, without any indications of inhibited mitochondrial energy production. Acute hypoxia induces increased respiratory capacities within 30-60 min. Under chronic conditions, alterations of mitochondrial cytochrome concentrations accompany the increased respiratory capacities. Instead of the decreased efficiency of mitochondrial energy-producing mechanisms induced by ischemia, hypoxia induces increased efficiency of energy production.


[Epidemiology of R plasmid and transferable antibiotic-resistance (author's transl)]. The AA. review the basic characteristics of extra-chromosomal antibiotic-resistance along with research guide-lines concerning the epidemiology of R plasmids. They then report the results of their investigations on isolated Salmonellae during the years 1971-1977. While S. typhimurium and S. wien showed an high incidence of transferable antibiotic-resistance, other Salmonellae belonging to 32 different serotypes had low incidence rates.


Parainfluenzavirus upper respiratory tract illnesses in partially immune adult human subjects: a study at an Antarctic station. Outbreaks of respiratory tract illnesses (RTI) in adult humans during October and November 1975 at McMurdo Station, Antarctica, were investigated by viral isolation and serologic procedures. The recovery of viral agents was enhanced by use of cell cultures in the field. Recoveries of parainfluenzaviruses types 1 and 3 and rhinoviruses were made from 10 of 39 nasal washings. Parainfluenzaviruses types 1 and 3 accounted for 50 and 30 per cent, respectively, of the total viruses recovered during October and November 1975. Acute and convalescent sera collected from 32 adult humans were examined for antiviral antibody by hemagglutination inhibition (HI) and radioimmunoassay (RIA) techniques. Serologic responses (HI and RIA) confirmed that parainfluenzaviruses were the predominent cause of RTI at McMurdo Station during that time. The temporal relationship between parainfluenzaviral diseases occurring in US communities and at McMurdo suggests that these viruses are transported to the Antarctic by personnel originating within the US. Standardization of the RIA allowed sequential assay of large numbers of sera using multiple preparations of radiolabeled indicator antibody, while ensuring the reproducibility of antiviral antibody titers to within one twofold dilution between subsequent labeled antibody preparations. The RIA detected both lower levels of virus specific antibody and more serologic responses than could be detected by HI.


Changes in intracellular cyclic AMP levels of human peripheral blood lymphocytes in bronchial asthma. Peripheral blood lymphocytes of asthmatic patients in the acute stage manifest low basal levels of cyclic AMP. These levels were higher in the lymphocytes of patients in remission than in controls. Trypsin treatment of the lymphocytes increased cyclic AMP content to almost the same additional activation site of the receptor in theophylline- and catecholamines-treated patients.


[CNV and SEP in shoe-industry workers affected by neuropathy due to toxic effects of adhesive solvents (author's transl)]. The sensitivity of the CNV and somatosensory evoked potentials (SEP) was assessed in shoe industry workers suffering from neurotoxic effects of adhesive solvents. We have examined 21 patients with clear electroneuromiographic and clinical signs of polyneuropathy as well as EEG signs of diffuse brain damage. 10 normal volunteers served as a control group. The maximal motor conduction velocity (MMCV) was considerably reduced in all patients. The maximal sensory conduction velocity (MSCV) was in the lower normal range (or borderline) in 12 patients, whereas in 9 or more severe decrement was detected. In comparison with normal subjects, none of the patients showed clear differences in latency or amplitude of SEP components (p always greater than .2). It was very easy to elicit CNVs over all areas explored and all the 10 patients showed normal characteristics. These results, therefore, suggest that CNV and SEP are not helpful for an early diagnosis of toxic effects of the solvents on the function of the central and peripheral nervous system.


[Müllerian papillary adeno-fibromata and müllerian adenosarcomata. Mülleromas and mixed malignant müllerian growths contingent with benign epithelium (author's transl)]. The authors present 6 cases of müllerian papillary adeno-fibromata (or mixed mesodermal müllerian benign tumours) of which 4 are anatomo-clinical presentations and 2 are histological presentations, as well as 3 cases of müllerian adenosarcoma (mixed müllerian mesodermal tumour with stroma only sarcomatous). The study of these cases added on to those found in the literature leads to the following conclusions: The first group can be accepted. The second group will be of mild malignancy. All the same, 4 of which the sarcomatous nature is not in doubt at all (malignant from the start) recurred rapidly. Those in which the sarcomatous nature was attenuated or doubtful (15 cases) seemed to have a much longer evolution with some local recurrences. Only 3 patients have been followed up for more than 5 years. It is absolutely essential to carry out further observations before confirming for certain the prognosis of this new type of tumour.


[Hormone receptors in breast cancer. Thoughts on the use of their determination as a test of hormone dependence in a hospital routine (author's transl)]. The determination of hormone receptors in tumour tissues including, among others, breast tumours is becoming routine examination in a greatly increasing number of clinical departments. We draw the attention of clinicians to the criteria of the quality necessary in such quantitative measurements and the limits of such an examination if it is to prove useful in directing the therapy of those clinics that use it in the light of the experience we have gained in the last 4 years.


[A thousand obstetrical sutures of the perineum using a slow absorption synthetic suture material (author's transl)]. A thread which is absorbed slowly and which has a polyglycolic acid base has been used since 1975 in the Sainte-Foy-les-Lyon Maternity Unit to sew up episiotomies and perineal tears. The authors had 98.70 p. 100 of excellent results out of 1082 cases from the anatomic, aesthetic and functional angle. This technique appears to be a useful one in early mobilisation in obstetrics.


[The effect of sodium polyanethol sulfonate (SPS) on the growth of bacteria in blood cultures]. The effect of SPS on growth of bacteria in blood cultures were studied. For that reason, 752 blood cultures (in media with and without SPS) were detected and found that SPS may help to growth only micrococcus species.


[Evaluation of the reflectometric method using the skin of the toad Bufo arenarum in vitro for the determination of plasma MSH]. The aim of the present paper was to investigate the following points, by using the reflectometric bioassay for melanocyte-stimulating hormone (MSH) with skin of the toad Bufo arenarum: (1) To study if solutions of alpha-MSH prepared in rat plasma and amphibian Ringer, or dilutions of plasma overloaded with endogenous MSH, could render in acceptable dose-response lines. 2) To investigate if ACTH at a very high level of plasma concentration could interfere in the bioassay 3) Observing if the storage of plasma could inactivate the inherent MSH activity. 4) Searching for the response of the skins along the 12 months of the year. The results showed that: Acceptable dose-response lines were obtained with the three types of dilutions and parallelism was found among the three lines. ACTH does not interfere in the assay. Almost a total inactivation of MSH was detected after storing the plasma at room temperature for 4 hours; Trasylol or low temperature preserved the hormone activity. Maximal response of the skin corresponded to the minimal concentration of 1 562 pgr/ml, during the whole year. Minimum detectable concentration varied within the range 24-390 pgr/ml, depending on the month when the study was performed.


Differential beta-adrenergic sensitivity of atrial tissue assessed by chronotropic and inotropic responses to several sympathomimetic amines. The relative inotropic and chronotropic activity of epinephrine, norepinephrine, isoproterenol, dopamine, dobutamine and ethylephrine was studied on isolated rat atria at 31 C. Dose-response curves were expressed as a percentage of the maximum response. Both atria were used for testing chronotropic effects. Inotropic dose-response curves were performed on left atria driven at 1 Hz, 1.6 Hz, 2.8 Hz and 6.6 Hz. Epinephrine and norepinephrine had a greater chronotropic action than their inotropic one throughout the range of stimulation employed. Their pD2-values changed from 8.12 +/- 0.06 and 8.05 +/- 0.05 to 7.19 +/- 0.03 and 7.14 +/- 0.05, respectively. Isoproterenol, dopamine and ethylephrine produced the same degree of chronotropic and inotropic stimulation. Dobutamine exerted the same effect on heart rate and developed tension at 1 Hz. pD2-values for dobutamine were significantly lower (p less than 0.01, p less than 0.001, p less than 0.001) when the frequency of stimulation was increased to 1.6, 2.8 and 6.6 Hz. These findings may be due to the influence of the negative staircase phenomenon, the temperature and the distribution of beta 1 and beta 2 receptors.


Protein quinone complexes. Bovine plasma albumin and halogenated p-quinones. A colloidal dispersion of chloranil in water or an aqueous solution of an amino acid shows an ESR signal characteristic of the semiquinone radical anion. The signal is broadened in the presence of bovine plasma albumin, and the available evidence supports the idea that the freedom of the free radical is restricted by a weak association with a specific site in the protein.


The effects of carbodiimides on functions associated with the energy-conservation mechanism in beef heart sub-mitochondrial particles. N,N'-di-n-propyl-, N,N'di-n-butyl-, N,N'-di-n-pentyl-, N,N'di-n-hexyl-, N,N'di-n-octoyl, N,N'-dibenzhydryl-, and N,N'-dibenzhydrylcarbodiimides were synthesized. They were all effective inhibitors (2 nmoles carbodiimide per milligram protein) of the ATP-driven reduction of NAD by succinate and the ATP-driven transhydrogenase activities catalyzed by beef heart submitochondrial particles (SMP). They had no effect on the nonenergy-linked transhydrogenase and stimulated the succinate-driven aerobic transhydrogenase activity of beef heart SMP. It was concluded that they exert their effects by reacting with the N,N'-dicyclohexylcarbodiimide-binding protein. Water-soluble carbodiimides were not effective inhibitors.


[Diagnosis and therapy of hepatocellular cancer]. A patient with hepatocellular carcinoma of the right liver lobe is presented by case report. Although a right side hemihepatectomy was performed, a recidive was detected 3 months later. Since then he received adriamycin-therapy and 14 months after operation, he is almost completely free of pain. Diagnosis, and surgical as well as internal therapy, are presented.


[Preliminary data on activation of the enzyme of the metabolism of phospholipid bases in various areas of the brain]. The scope of this work was to study the effect of imydazole on base-exchange enzymic system in selected cerebral areas. We have previously demonstrated that imydazole was an activator of phosphatidylserine synthesis in slices of caudate nucleus. This effect lacked in the omogenate we had supposed that this activation by imydazole was not directed on base-exchange enzymic system, but was induced by decrease of cellular cyclic nucleotides amount. Therefore we have investigated the effect of dibutirril-AMPciclic in selected cerebral areas. In addition it was useful to state if activation by imydazole was area-specific or not.


[Preliminary data on the behavior of the enzyme of metabolism of phospholipid bases in human tumor preparations]. The A.A. have investigated the phospholipids base-exchange enzyme system in the solid tumours in order to state if a correlation between this activity and the variation of the cellular ciclic nucleotides amount was possible considering that these compounds have been reported to undergo a variation in tumour compared with normal tissues. They report some previous results in a lung tumour and in an endometrial carcinoma, were they have found a big increase in the PhS synthesis. Such increase was possible to be seen in the endometrial carcinoma only after a stimulation by 17-beta-estradiol and it was reversed in this case by doxorubicine. These results suggest that an alteration of the PhS synthesis should be one of numerous peculiarity of the neoplastic cell.


[Changes in phosphatidylserine synthesis in human rectal tumors]. We have investigated the phosphatidylserine metabolism in human adenocarcinoma. This phospholipid is involved in the regulation of several enzymic activities of plasma membranes; therefore it could be useful to state if some alteration of tumour membranes propriety depend on variation of phosphatidylserine synthesis. Our results make evident that in such human tumours the phosphatidylserine synthesis is greatly augmented.


Protective effects of rat splenic lymphocytes and peritoneal exudate cells on herpes simplex virus infection of rat dorsal root ganglia in culture. An in vitro model system was developed to study the effects that immune cells might have on herpes simplex virus (HSV) infection of rat dorsal root ganglia in culture. Our results demonstrate that rat splenic lymphocytes and peritoneal exudate cells are incapable of replicating HSV even if these cells come from sensitized animals and are stimulated in vitro. Both cell preparations can offer some protection to rat dorsal root ganglia from the effects of HSV infection. The data suggest that an immunologically non-specific (not mediated by sensitized cells) type of protection is important to neurons, while an immunologically specific (mediated by sensitized cells) protection is most beneficial to fibroblasts. This system can be utilized to study the mechanism of latency since the neurons of sensory ganglia are the natural site of latent herpes virus.


An accountability model for integrating information systems, evaluation mechanisms, and decision making processes in alcohol and drug abuse agencies. This article has attempted to demonstrate that decision making and evaluation can be carried out in a systematic fashion only if agencies make a commitment to do so, and only if adequate systems are established. The management information system is the most expensive and most sophisticated component of the integrated model presented here. Its existence, in some fashion, is essential to the operation of the model. Contrary to what many managers may believe and practice, the management information system is not in itself the final solution to evaluation. Neither is the evaluation a panacea for all program ills. Evaluation can provide the information required to meet the ever increasing demands for agency or program accountability evaluation can also provide insights for future decisions to change or alter the allocation of resources. Such evaluation must be carefully planned and implemented; and, at the state level, can be successful only if executed in a systematic manner as suggested here. Regardless of the degree of sophistication of any system, it will work only when supported by users in the local treatment centers. If the model employed does little to serve them, it is not a model worth considering. It is with these needs in mind that this model was developed.


The dynamics of K+ leakage and recovery in cerebral ischemia. A combination of K+/DC surface electrode and a fiberoptic fluorometric probe are applied to measurements in brain during cerebral ischemia. The kinetics of the responses of extracellular K+ activity and intracellular NADH fluorescence in the gerbil cerebrum following reversible carotid ligation are measured. K+e shows a two-phase response to carotid occlusion and an extended recovery phase following recirculation. The length of the recovery phase is dependent on the duration and severity of the ischemic period. In the gerbil model the degree of communication in the anterior circulation is variable, whereas a bilateral carotid occlusion is presumed to give complete cerebral ischemia. Pyridine nucleotide fluorescence serves as an indicator of the degree of ischemia. Bilateral carotid occlusions of up to 35 minutes in duration were performed. K+e reaches 30--50 mEq/liter in the extracellular space within the first two minutes. This represents cell depolarization and equilibration of K+ activity levels. Recovery appears to be complete in terms of the ability of the system to clear raised levels of K+e from the extracellular space.


Evaluation of a new vaginal contraceptive. A new chemical contraceptive in the form of a vaginal suppository was evaluated in a series of in vitro and in vivo tests and compared to a leading contraceptive foam and cream. Good in vitro spermicidal effect was observed, but problems in solubility and the position of the suppository against the external cervical os may lead to less clinical effectiveness if not properly evaluated prior to its use.


Long-term effects of metaproterenol in asthmatic children. Oral metaproterenol was administered daily for 3 months to asthmatic children. The study was designed to determine whether chronic tolerance developed to this drug. Initial and final crossovers on the first 2 and the last 2 days of the investigation to a test dose of metaproterenol and placebo failed to show tolerance as indicated by improvement in lung volumes and dynamic mechanics of breathing after metaproterenol. Forced expiratory volume in 1 sec was the test most consistent in demonstrating bronchodilation; the action of metaproterenol. Forced expiratory volume in 1 sec was the test most consistent in demonstrating bronchodilation; the action of metaproterenol appeared to last at least 5 hours as measured by this test. T of distribution of ventilation, e.g., single- and multiple-breath nitrogen washout tests, were not consistently altered by metaproterenol. These tests did not appear to be sufficiently sensitive to detect improvement even when airway resistance decreased and forced expiratory volume in 1 sec increased toward the normal range.


The synthesis and metabolism of the lipoproteins implicated in atherosclerosis. The basic concepts of lipoprotein biogenesis and metabolism have been briefly reviewed. Emphasis has been placed upon the distinctive roles of the liver and intestine in apolipoprotein synthesis and delivery, upon the complex interchange of lipids and apolipoproteins between lipoproteins which accompanies nascent lipoprotein maturation and metabolism within the vascular compartment, upon the primary role of these latter processes in the genesis of low density lipoprotein and upon remodelling of nascent lipoproteins to achieve a structure suitable for the function of the mature between the liver, intestine and peripheral tissues.


Relationship between Na+-dependent respiration and Na+ + K+-adenosine triphosphatase activity in the action of thyroid hormone on rat jejunal mucosa. Administration of three successive doses of triiodothyronine (T3) (50 micrograms/100 g body wt), given on alternate days to thyroidectomized and euthyroid rats, stimulated oxygen consumption (QO2) and Na+ transport-dependent respiration (QO2 [5]) in the stripped jejunal mucosa, a preparation that consisted mostly of epithelial cells. The increase in QO2(t) accounted for 57% of the increment in QO2 in the transition from the hypothyroid to the euthyroid state and for 29% of the increment in the transition from the euthyroid to the hyperthyroid state. Administration of T3 to hypothyroid rats also increased the yield of epithelial cells. Injection of T3 into thyroidectomized and euthyroid rats increased the specific activity (at Vmax) of the (Na+ + K+)-dependent adenosine triphosphatase (NaK-ATPase) in jejunal crude membrane preparations. No significant change was recorded in the activity of Mg-ATPase in the same preparation. The ratio of QO2/NaK-ATPase and QO2(t)/NaK-ATPase in the various thyroid states remained constant, indicating proportionate increased in the respiratory and enzymatic indices. The effect of administration of T3 to thyroidectomized rats on the number of NaK-ATPase units (recovered in the crude membrane preparation) was estimated by: (a) Na+ + Mg++ + ATP-dependent binding of [3H]-ouabain to crude membrane fractions, and (b) the amount of the phosphorylated intermediate formed in the NaK-ATPase reaction from AT32P(gamma). Estimates were obtained of the maximal number of [3H]ouabain binding sites (Nm) and dissociation constants (Kd). Nm for [3H]ouabain and Nak-ATPase specific activity increased to about the same extent after T3 administration to thyroidectomized rats, with no change in the apparent Kd values. The amount of phosphorylated intermediate formed in jejunal crude membrane preparations also increased significantly. Thus, thyroid hormone administration may increase the number of active Na+pump sites in the plasma membrane. The apparent increase in the number of Na+ pump sites also correlated with the hormone dependent increases in QO2 and QO2(t).


Interactions between quaternary lidocaine, the sodium channel gates, and tetrodotoxin. A voltage clamp technique was used to study sodium currents and gating currents in squid axons internally perfused with the membrane impermeant sodium channel blocker, QX-314. Block by QX-314 is strongly and reversibly enhanced if a train of depolarizing pulses precedes the measurement. The depolarization-induced block is antagonized by external sodium. This antagonism provides evidence that the blocking site for the drug lies inside the channel. Depolarization-induced block of sodium current by QX-314 is accompanied by nearly twofold reduction in gating charge movement. This reduction does not add to a depolarization-induced immobilization of gating charge normally present and believed to be associated with inactivation of sodium channels. Failure to act additively suggests that both, inactivation and QX-314, affect the same component of gating charge movement. Judged from gating current measurement, a drug-blocked channel is an inactivated channel. In the presence of external tetrodotoxin and internal QX-314, gating charge movement is always half its normal size regardless of conditioning, as it QX-314 is then permanently present in the channel.


Intrinsic electric fields and proton diffusion in immobilized protein membranes. Effects of electrolytes and buffers. We present a theory for proton diffusion through an immobilized protein membrane perfused with an electrolyte and a buffer. Using a Nernst-Planck equation for each species and assuming local charge neutrality, we obtain two coupled nonlinear diffusion equations with new diffusion coefficients dependent on the concentration of all species, the diffusion constants or mobilities of the buffers and salts, the pH-derivative of the titration curves of the mobile buffer and the immobilized protein, and the derivative with respect to ionic strength of the protein titration curve. Transient time scales are locally pH-dependent because of protonation-deprotonation reactions with the fixed protein and are ionic strength-dependent because salts provide charge carriers to shield internal electric fields. Intrinsic electric fields arise proportional to the gradient of an "effective" charge concentration. The field may reverse locally if buffer concentrations are large (greater to or equal to 0.1 M) and if the diffusivity of the electrolyte species is sufficiently small. The "ideal" electrolyte case (where each species has the same diffusivity) reduces to a simple form. We apply these theoretical considerations to membranes composed of papain and bovine serum albumin (BSA) and show that intrinsic electric fields greatly enhance the mobility of protons when the ionic strength of the salts is smaller than 0.1 M. These results are consistent with experiments where pH changes are observed to depend strongly on buffer, salt, and proton concentrations in baths adjacent to the membranes.


Block of sodium conductance and gating current in squid giant axons poisoned with quaternary strychnine. Quaternary strychnine blocks sodium channels from the axoplasmic side, probably by insertion into the inner channel mouth. Block is strongly voltage dependent, being more pronounced in depolarized than in resting axons. Using potential steps as a means to modulate the level of block, we investigate strychnine effects on sodium and gating currents at +50 and -50 mV. We analyze our data in terms of the simplest possible model, wherein only an open channel may receive and retain a strychnine molecule. Our main findings are (a) block by strychnine and inactivation resemble each other and (b) block of sodium and gating currents by strychnine happen with closely similar time-courses. Our data support the hypothesis of Armstrong and Bezanilla (1977) wherein an endogenous blocking particle causes inactivation by inserting itself into the inner mouth of the sodium channel. Quaternary strychnine may act as an artificial substitute for the hypothetical endogenous blocking particle. Further, we suggest that at least 90% of the rapid asymmetrical displacement current in squid axons is sodium channel gating current, inasmuch as quaternary strychnine can block 90% of the displacement current simultaneously with sodium current.


Proton T1 study of coverage parameter changes in tissues from tumor-bearing mice. measurements of water proton spin-lattice relaxation time, T1, at 20 and -15 degrees C have been performed in spleen, kidney, liver, and muscle tissues from tumor-bearing mice, as well as in tumors grown in their dorsal subcutaneous tissues. All mice used were either from the C3H/HeJ or BALB/c strain. At - 15 degrees C the T1's of tissues of a given type from tumor-bearing and healthy mice are essentially the same. It is shown that in spleen the increased T1 from tumor-bearing mice can only be explained in terms of a large change in the water coverage parameter of macromolecules. In liver, muscle, and tumors the increased water content accounts for the changes in T1, while kidney represents an intermediate case.


Photodynamic induction of an oncogenic virus in vitro. Infectious simian virus 40 (SV40) was induced from SV40-transformed hamster kidney cells by treatment with proflavine and visible fluorescent light. The optimum levels of SV40 induced were about three orders of magnitude above spontaneous background levels observed with untreated cells. No virus induction above background levels was found by treatment of cells with either proflavine or light alone.


Development of electron spin polarization in photosynthetic electron transfer by the radical pair mechanism. We have extended the radical pair theory to treat systems of membrane-bound radicals with g tensor anisotropy. Analysis of the polarized electron paramagnetic resonance (EPR) signals of P700+, originating from photosystem I of higher plants, in terms of the radical pair mechanism provides information about the sequence of early electron acceptors. To account for the orientation dependence of the line shape and integrated area of this polarized signal, we propose the electron transfer sequence to be P700 leads to A1 leads to X leads to Fd(A, B), where A1 is a small organic molecule (possibly chlorophyll), X is the acceptor species observed recently in low-temperature EPR studies, and Fd(A, B) are the ferredoxin iron-sulfur centers A and B. Our calculations provide information about the life-times of A1-, and X-, and their exchange interactions with P700+. We also find supporting evidence for the orientation of X- in the thylakoid membrane reported recently by G. C. Dismukes and K. Sauer (Biochim. Biophys. Acta. 504:431-445.).


Crystalline state disorder and hyperfine component line widths in ferric hemoglobin chains. In X-band electron paramagnetic resonance spectra from single crystals of horse ferric hemoglobin, observed line widths at the low- and high-field extrema are 30 and 24 g, and as much as 400 G in the intermediate region. This behavior is similar to that of ferric myoglobin. Due to large anisotropy in the g-tensors, the line width variation can be accounted for on the basis of heme orientation disorder. This disorder is characterized by an angle, determined here by two independent methods. In these computations Gaussian disorder on a sphere is assumed. The disorder angle is found to be constant on the sphere and about 4 degrees for both alpha- and beta- chains. Treatment of crystals with heavy water (buffer) increases the disorder. Since ligand nitrogen hyperfine couplings are available from hemoglobin electron nuclear double resonance, single crystal electron paramagnetic resonance spectra can be simulated by superimposing hyperfine bands, where the line width of the component bands is a variable and the disorder model above is employed. Comparison with observed resonances fixes the hyperfine component line widths. These component line widths from ferric hemoglobin in the crystalline state are found to be smaller than those in frozen solution.


Quantitative evaluation of contributions to electron paramagnetic resonance line widths in ferric hemoglobin single crystals. The contributions to the dipolar broadening of ferric magnetic resonances, from crystals of hemoglobin for which the atomic coordinates are known, have been calculated. The total second moment of the g = 2 resonance so determined is about 50 (MHz)2 or 5.0 G (peak-to-trough), figures consistent with the range of values found from analysis of experimental data. Two-thirds of this second moment comes from the two protons of the H2O molecule coordinated to the iron. Treatment with D2O is predicted to reduce the total second moment at g = 2 to about 25 (MHz)2, whereas the experimental measurements on single crystals show no decrease. If the structure of the tetramer is assumed to be the same when in solution as in the crystal, the total second moment is readily redetermined for hemoglobin in solution; the value so obtained is found to be significantly smaller than that from analysis of the g = 2 resonance measured in frozen solution. These two unexpected observations can be explained in terms of distributions in spin Hamiltonian parameters, the spread depending upon the nature of the sample--crystal or solution, ordinary or heavy water-treated. This distribution in H2O and D2O solutions appears to be about the same, since the measured differences in component line width agree with the calculated difference in dipolar contributions.


n-Alkanols potentiate sodium channel inactivation in squid giant axons. The effects of n-octanol and n-decanol on nerve membrane sodium channels were examined in internally perfused, voltage-clamped squid giant axons. Both n-octanol and n-decanol almost completely eliminated the residual sodium conductance at the end of 8-ms voltage steps. In contrast, peak sodium conductance was only partially reduced. This block of peak and residual sodium conductance was very reversible and seen with both internal and external alkanol application. The differential sensitivity of peak and residual conductance to alkanol treatment was eliminated after internal pronase treatment, suggesting that n-octanol and n-decanol enhance the normal inactivation mechanism rather than directly blocking channels in a time-dependent manner.


Magnetic resonance studies of the spatial arrangement of glucose-6-phosphate and chromium (III)-adenosine diphosphate at the catalytic site of hexokinase. The interaction of CrADP, an exchange-inert paramagnetic analogue of Mg-ADP, with yeast hexokinase has been studied by measuring the effects of CrADP on the longitudinal nuclear relaxation rate (1/T1) of the protons of water and the protons and phosphorus atom of enzyme-bound glucose-6-P. The paramagnetic effect of CrADP on 1/T1 of water protons is enhanced upon complexation with the enzyme. Titrations measuring this paramagnetic effect at several enzyme concentrations in the presence of glucose-6-P yielded a characteristic enhancement factor for 1/T1 of water protons and the dissociation constant of CrADP from the ternary enzyme . ADPCr . glucose-6-P complex. The latter value (2 mM) is similar to that obtained from kinetic inhibition studies (Danenberg and Cleland [1975]. Biochemistry. 14:28). The presence of glucose-6-P increased the enhancement of the water relaxation rate by enzyme-bound CrADP, suggesting the formation of an enzyme . CrADP . glucose-6-P complex. The existence of such a complex was confirmed by the observation of a paramagnetic effect of enzyme-bound CrADP on the l/T1 of the 31P-nucleus and protons of enzyme-bound glucose-6-P. From the paramagnetic effects of enzyme-bound CrADP on the relaxation rates of the 31P-nucleus and the carbon-bound protons of glucose-6-P in the enzyme . ADPCr . glucose-6-P complex, using the correlation time of approximately 0.7 ns, determined from the magnetic field-dependence of 1/T1 of water protons over the range 24.3-360 MHz, a Cr3+ to phosphorus distance of 6.6 +/- 0.7 A and Cr3+ to alpha- and beta-anomeric proton distances of 8.9 and 9.7 A were calculated. These results imply the absence of a direct coordination of the phosphoryl group of glucose-6-P by the nucleotide-bound metal on hexokinase but indicate van der Waals contact between a phosphoryl oxygen of glucose-6-P and the hydration sphere of the nucleotide-bound metal. The distances are consistent with a model that assumes molecular contact between the phosphorus of glucose-6-P and a beta-phosphoryl oxygen of ADP suggesting an associative phosphoryl transfer. Because after phosphorylation of ADP, the metal ion is coordinated to the transferred phosphoryl group, the overall migration of the phosphoryl group during the phosphoryl transfer is approximately 3.6 A toward the nucleotide-bound metal. Little or no catalysis of phosphoryl transfer from glucose-6-P to alpha, beta-bidentate or beta-monodentate CrADP ( less than or equal to 0.05% of the rate found with MgADP) occurred in the presence of hexokinase, as monitored by glucose formation in a coupled assay system using glucose oxidase and peroxidase. The ability of beta, gamma-bidentate CrATP to act as a substrate (Danenberg and Cleland [1975].


Electrolyte transport across a simple epithelium. Steady-state and transient analysis. A simple transporting epithelium is represented as a cellular compartment, compliant in all dimensions, and a paracellular channel, of arbitrary shape, between well-stirred mucosal ans serosal baths. The equations for mass balance, Poiseuille flow, and the Nernst-Planck equation are used to describe the continuous behavior of the system along cell and channel, whereas passive transport across membranes is given by the relations of Kedem and Katchalsky. Time-dependent terms are retained to permit study of transient phenomena. Boundary conditions at the baths demand only mass conservation and specify no a priori estimates of the system variables. A numerical model containing Na+,K+,Cl-, and impermeant cellular anions is formulated with membrane parameters taken from the literature on Necturus gallbladder. The differential equations are represented as a finite difference scheme and solved using Newton's method. It appears that apical cellular NaCl cotransport is necessary to obtain a reasonable cell chloride concentration. Investigation of the osmolality of the transepithelial flow shows that at steady state a leaky epithelium cannot separate baths of substantially different tonicity, although this does not guarantee isotonic transport between equiosmolar media. Changes in bath pressure, application of transepithelial electrical potential, and simulation of ion-substitution experiments are performed to understand the role of membrane permeabilities in determining the dynamic behavior of the epithelium.


Modifications of sodium channel gating in Myxicola giant axons by deuterium oxide, temperature, and internal cations. In dialyzed Myxicola axons substitution of heavy water (D2O) externally and internally slows both sodium and potassium kinetics and decreases the maximum conductances. Furthermore, this effect is strongly temperature dependent, the magnitude of the slowing produced by D2O substitution decreasing with increasing temperature over the range 3-14 degrees C with a Q10 of approximately 0.71. The relatively small magnitude of the D2O effect, combined with its strong temperature dependence, suggests that the rate limiting process producing a conducting channel involves appreciable local changes in solvent structure. Maximum conductances in the presence of D2O were decreased by approximately 30%, while the voltage dependences of both gNa and gK were not appreciably changed. In contrast to the effects of heavy water substitution on the ionic currents, membrane asymmetry currents were not altered by D2O, suggesting that gating charge movement may preceed by several steps the final transformation of the Na+ channel to a conducting state. In Myxicola axons the effect of temperature alone on asymmetry current kinetics can be well described via a simple temporal expansion equivalent to a Q10 of 2.2, which is somewhat less than the Q10 of GNa activation. The integral of membrane asymmetry current, representing maximum charge movement, is however not appreciably altered by temperature.


Conditioning hyperpolarization-induced delays in the potassium channels of myelinated nerve. Hyperpolarizing conditioning pulses delay the onset of potassium channel current in voltage-clamped myelinated nerve fibers. Both the development of and recovery from this conditioning are approximately exponential functions of time: the time constants are functions of the conditioning voltage. The delay is larger and develops faster for more hyperpolarized conditioning pulses. The magnitude of the delay (but not the rate of development or recovery) depends upon the test potential-small test depolarizations produce larger delays than large depolarizations. The currents with and without the conditioning pulse cannot be made to superimpose by a simple time translation.


Structure of the replication fork in ultraviolet light-irradiated human cells. The DNA extracted from xeroderma pigmentosum human fibroblasts previously irradiated with 12.5 J/m2 of UV light and pulse-labeled for 45 min with radioactive and (or) heavy precursors, was used to determine the structural characteristics of the replication fork. Density equilibrium centrifugation experiments showed that a fork moved 6 micrometer in 45 min and bypassed 3 pyrimidine dimers in both strands. The same length was covered in 15-20 min in control cells. The delay in irradiated cells was apparently due to pyrimidine dimers acting as temporary blocks to the fork movement. Evidence for this interpretation comes from kinetics of incorporation of [3H]thymidine into DNA, which show that the time necessary to attain a new stable level of DNA synthesis in irradiated cells is equivalent to that required for the replication fork to cover the interdimer distance in one strand. On the other hand, the action of S1 nuclease on DNA synthesized soon after irradiation gives rise to a bimodal distribution in neutral sucrose gradients, one peak corresponding to 43 X 10(6) daltons and the other to 3 X 10(6) daltons. These two DNA species are generated by the attack of the S1 nuclease on single-stranded regions associated with the replication fork. A possible explanation for these results is given by a model according to which there is a delayed bypass of the dimer in the leading strand and the appearance of gaps opposite pyrimidine dimers in the lagging strand, as a direct consequence of the discontinuous mode of DNA replication. In terms of the model, the DNA of 43 X 10(6) daltons corresponds to the leading strand, linked to the unreplicated branch of the forks, whereas the piece of 3 X 10(6) daltons is the intergap DNA coming from the lagging strand. Pulse and chase experiments reveal that the low molecular weight DNA grows in a pattern that suggests that more than one gap may be formed per replication fork.


Augmentation of the in vitro humoral immune response by pharmacologic agents. II: comparison of the effects of antiproliferative agents with DBcAMP. We have compared the stimulatory activity of DBcAMP with various antiproliferative agents on the induction of the humoral immune response. When they are present only during an early stage of immune induction, DBcAMP, colcemid, cytosine-arabinoside, hydroxy urea, and high specific activity 3H-thymidine can all enhance the primary 19s antibody response to SRBC. In contrast, each of these agents inhibits the PFC response, when they are incubated with the cells during late stages of induction of humoral immunity. Because all of these agents can inhibit proliferation of cultured cells, the results suggested that DBcAMP and other agents that elevate cAMP could augment humoral immunity via their effects on cellular proliferation. However, we also found that although each agent could modulate induction of the immune response to SRBC, only DBcAMP produced a dose- and time-dependent augmentation of the response to DLF. We conclude that although antiproliferative effects of drugs may contribute to augmentation of some humoral antibody responses, this effect alone is insufficient to account for the mechanism by which agents that elevate intracellular levels of cAMP produce enhancement of humoral immunity.


Cyclic nucleotide changes in murine lymphocytes following thymosin incubation in vitro. Thymosin fraction 5 and a more purified acidic fraction of thymosin at a 100-fold lower concentration elevated cGMP levels, but not cAMP levels, in murine thymocytes. When both thymus and spleen lymphocytes were fractionated via a BSA gradient procedure, both cAMP and cGMP basal values varied depending on the density of the subpopulations. Thymosin Fr5 did not elevate cAMP in any thymus subpopulation of lymphocytes obtained from the BSA density gradients. The cGMP elevation due to thymosin Fr5 in thymocytes was maximal in the most buoyant thymocytes, and no elevation of cGMP was detected in nude mouse spleen lymphocytes. These results suggest that the cGMP elevation may be an early event in the thymosin-mediated differentiation of a more mature subpopulation of thymocytes. They also suggest that utilization of subpopulations may be necessary for the complete determination of the effects of agents on cyclic nucleotide values of lymphocytes.


Adriamycin-induced antitumor response in lethally irradiated mice. Lethally irradiated mice pretreated with a wide range of Adriamycin (ADM) doses were tested for their in vivo antitumor response against transplanted lymphoma cells. Tumor growth, as assessed by 125IUdR uptake, was markedly impaired by treatment with ADM by a variety of different administration schedules. This ADM-induced antitumor response was largely dose-dependent, occurred regardless of route of administration, and was detectable as late as 15-30 days following drug treatment. No genetic restriction could be found to regulate the development of this response, since tumor growth inhibition occurred in syngeneic as well as allogeneic tumor-host combinations. ADM-induced antitumor response did not appear to be due to direct antitumor action by the drug, but rather to some interaction with host immune mechanisms. Antimacrophage agents, such as silica or carrageenan, abrogated the response. The possible implication of different cells as effectors of this response is discussed.


Increased polymorphonuclear leukocyte cGMP levels induced by the human lympholine, leukocyte migration inhibitory factor (LIF). The possible involvement in vitro of 3', t'-cyclic GMP (cGMP) in the mechanism of action of the lymphokine, leukocyte migration inhibitory factor (LIF), was investigated. Partially purified LIF-rich supernatants, but not their control counterparts, induced a 2-fold increase in the cGMP levels of purified human polymorphonuclear (NMN) leukocytes. The effect was no influenced by heat-inactivated horse serum; it was manifested within 3 min of exposure to LIF and it subsided within 180 min. LIF and the supernatant factor responsible for the cGMP-generating effect were both rendered inactive by treatment with the serine esterase and protease inhibitor, phenylmethylsulfonyl fluoride, indicating that these factors are closely related, if not identical. A potent phosphodiesterase inhibitor, dipyridamole (2 x 10(-4) M), induced a 3- to 5-fold increase in PMN leukocyte cGMP levels, but combined treatment with purified LIF and dipyridamole did not add to this effect. This suggests that the cGMP-generating factor acts on the biochemical pathway that degrades cGMP.


The effect of decreased growth rate on frequency and severity of osteochondrosis in pigs. Rapid weight gain seems to be related to high incidence and severity of osteochondrosis. For this reason the effect of weight gain on osteochondrosis was investigated. In one experiment including 18 pigs, frequency and severity of osteochondrosis in pigs on a low caloric intake and in those on a high caloric intake (ad libitum feeding) were compared. In another experiment 8 pigs from 2 litters of pigs with a mainly wild hog ancestry were studied. These pigs had a much lower growth rate than modern domestic pigs and their constitution was different. The results of these two experiments were unequivocal. Animals with a low weight gain, nutritionally or genetically induced, had an incidence of osteochondrosis which was close to zero. It was considered that the very high growth rate in the modern domestic pig is the main reason for the high incidence of osteochondrosis. High growth rate is the result of selective breeding and intensive feeding. There are some morphologic characteristics of the modern domestic pig similar to those seen in acromegaly in man and in animals treated with growth hormone.


Heredity of osteochondrosis, body constitution and leg weakness in the pig. A correlative investigation using progeny testing. Sixty-three boars of the Swedish Landrace and the Yorkshire breed were progeny tested. Production variables were correlated with frequency of osteochondrotic lesions at three sites, length of radius, and presence of leg weakness in 416 offspring. Osteochondrotic lesions in distal femur and humerus were found to be correlated with desirable production variables. No correlation existed between presence of osteochondrosis and leg weakness. Analysis of variance showed that breed and boar within breed influenced frequency and distribution of osteochondrosis. The pigs of the Yorkshire breed were less affected. Heritability estimates ranged from 0.14 to 0.30 for the osteochondrotic lesions at three sites. It should be noted that these figures are preliminary because of the limited number of offspring. It was concluded that a possibility exists to lower frequency of osteochondrosis in the pig with only some loss of valuable production qualities.


Osteochondrosis in the horse. I. A clinical and radiologic investigation of osteochondritis dissecans of the knee and hock joint. The clinical and radiologic features of osteochondritis dissecans in the knee and hock joint of horses are described. The material includes 91 horses, of which 43 had the lesion in one or both knee joints, and 48 in one or both hock joints. It was found that osteochondritis dissecans of the knee joint was more common than the one in the hock joint in thoroughbreds and halfbreds, while the lesion in the hock joint was most common in standardbred trotters. In the knee the lesion was most often located to the lateral trochlear ridge. In the hock joint the predilection site was the intermediate ridge of the tibia. The lesions were of obvious clinical importance and the knee joint was usually more severely affected than the hock joint. Prognosis was also more guarded in horses with knee joint involvement, regardless whether or not surgical treatment was instituted. Arthrotomy and removal of fragments in the osteochondral defects or of loose bodies was the standard surgical procedure. It was felt that this treatment improved prognosis somewhat, particularly in the horses with hock joint lesions. It was found that there are many similarities between osteochondritis dissecans in horses and in pigs and dogs. However, further investigation is needed to shed more light on etiology and pathogenesis and on the nature of osteochondritis dissecans in the horse.


Growth and skeletal development of the pig. Growth and skeletal development of the domestic pig (Swedish landrace and Yorkshire) are reported and the weight curve of males from birth to maturity included. Other parameters were tooth development and growth of certain bones. It was concluded that daily weight gain increases rapidly to an age of about 5 months. Sexual maturity is reached by both the male and female pig at about 5--6 months of age. At this time there is an inflection point on the weight curve. The period from 5--6 months to about 18 months of age is called adolescence. After 18 months of age the weight curve is flattened. The data from the domestic pigs were compared with the corresponding data of the wild European hog. It was found that the wild hog has a much slower weight gain.


Osteochondrosis in the horse. II. Pathology. An investigation was made of the pathology of osteochondritis dissecans of young foals and horses with clinical signs of the lesion. A randomly selected material of fetuses and young foals without clinical signs was also examined. It was demonstrated that osteochondritis dissecans is primarily a cartilaginous disease, as previously described in pigs and dogs. Thickening, disturbance of endochondral ossification, degeneration and necrosis of the cartilage were the four main features of osteochondritis dissecans. Cracks and fissures occurred in the degenerated and necrotic parts of the cartilage. This led to formation of cartilage flaps and eventually to loose bodies. It was shown that small pieces of subchondral bone could be ripped off when a cartilage flap was formed. This was one explanation as to why many flaps and loose bodies contained bone in contrast to the findings in pigs and dogs. Endochondral ossification could also take place in the thickened joint cartilage in some cases. Even some loose bodies could undergo endochondral ossification if they were well nourished. Osteochondritis dissecans was often found bilaterally in the knee and hock joint and this was interpreted as an indication that osteochondritis dissecans is a manifestation of a generalized condition called osteochondrosis. Simultaneous occurrence of lesions in joints other than the knee and hock and in several metaphyseal growth plates was another indication of the generalized nature.


Osteochondrosis in growing bulls. Pathology, frequency and severity on different feedings. Lesions typical for osteochondrosis were found in the joint cartilage and metaphyseal growth plates of growing bulls, of which many showed clinical signs of so-called leg weakness. Of 48 bulls included in the investigation, 23 were on intensive feeding and they all were found to have osteochondrosis at several locations. Of the 25 bulls of lower intensity feeding, only 13 had osteochondrosis. The lesions in those bulls were less severe than in the bulls on the high intensity feeding. At slaughter, when the lesions were inspected, the bulls on high energy feeding were much younger than the ones on the lower energy feeding. This must be kept in mind, when a comparison is made. As the bulls of the high intensity feeding were growing more rapidly, frequency and severity of osteochondrosis seemed to be correlated with rapid growth. Other etiological factors were, however, considered. A comparatively low intake of calcium in relation to daily weight increase may be one explanation for the high frequency in the bulls in high intensity feeding. Lack of exercise and large amounts of concentrates were possible explanations for the overall high frequency of osteochondrosis in the present material. The lesions in the joints and growth plates in the bulls were similar to those seen in pigs and horses. The reason why some partly or completely detached bodies contained bone is discussed. A possible connection between osteochondrosis and so-called subchondral bone cysts, as previously described in horses, is mentioned.


Tibial dyschondroplasia (osteochondrosis) in the turkey. A morphologic investigation. Sixty-seven Broadbreasted White Turkeys, 4 to 137 days old, were included in the investigation. Some were normal, while others were limping, and some had bowed legs. Immediately after destruction of the birds, the arteries of the legs were injected with Colorpaque, using a technique introduced by the author. The proximal part of one or both tibiae were examined radiographically, macroscopically, and microscopically. The microscopic examination included histology, microangiography, and microradiography. The normal development and morphology of the proximal tibia is presented. Thereafter an account is given of development and morphology of the tibia with dyschondroplastic lesions (retained cartilages). It was found that 39 of the 67 birds had some degree of tibial dyschondroplasia. The retained cartilage was seen in the posteromedial part of the proximal tibial metaphysis. The growth plate both of the normal and abnormal birds consisted of 3 layers of cells in order from the epiphysis--the one of proliferating cells, the one of transitional cells, and the one of hypertrophied cells. A main feature in tibial dyschondroplasia is accumulation of transitional cells, which do not have a vascular supply. By microangiographic means it was possible to demonstrate that the retained cartilage is surrounded by numerous vessels, which apparently cannot get into the cartilage. The reason for the loss of normal differentiation of the cartilage cells and concomitant disturbance of the endochondral ossification is obscure. It is speculated that changes on an ultrastructural and biochemical level are responsible for the pathologic condition. When the retained cartilage reaches a certain size, degenerative changes, necrosis, and fissures occur. Complete healing of the lesion was not seen in any of the birds, but partial disappearance of the proximal or distal part of the retained cartilage took place in some birds. It was concluded that the present investigation has shed light on the development and morphology of tibial dyschondroplasia, but it has also brought forward a number of questions about etiology. There seems to be good reason to believe that dyschondroplasia in the turkey is of similar nature as dyschondroplasia in broilers and osteochondrosis in domestic mammals.


Skeletal lesions in the broiler, with special reference to dyschondroplasia (osteochondrosis). Pathology, frequency and clinical significance in two strains of birds on high and low energy feed. The material consisted of 2,950 broilers of the variety Hybro Compact of two lines. An equal number of birds of both lines were given either a high energy feed (H) or a low energy feed (L), containing all nutrients known to be required by broilers. During the first 3 weeks, H and L starter feeds were given and thereafter H and L finishing feeds. At 21, 35, and 45 days of age, the birds were weighed in groups, and feed conversion calculated. At 21 days of age, the frequency of crooked toes and swollen hocks was registered. Birds were taken from each group for necropsy at regular intervals. Both clinically normal birds and those with locomotor disturbances were selected for necropsy, including radiographic examination. With the exception of 240 birds, which were kept for a long-term study, the birds on the H feed were slaughtered at 42 days of age and the ones on the L feed at 49 days of age. The long-term birds were slaughtered at regular intervals, weighed, and necropsied. The last birds were slaughtered at 134 days of age. A large number of skeletal lesions were found. They were: Twisted legs, slipped tendons, crooked toes, bowing of the proximal tibia, dyschondroplasia at different sites, fracture of the fibula, deformity of the spine, deviated sternum (with breast blisters), arthritis, and osteomyelitis. Leg weakness was found to be almost synonymous with skeletal deformities. Other causes were rare. The normal development and morphology of the skeleton and the morphology of tibial dyschondroplasia were the same as those previously described in the turkey. Dyschondroplasia was found not only in the proximal tibia but also in the distal tibia, proximal tarsometatarsus, proximal and distal femur, and to some extent also in the costochondral junction. Tibial dyschondroplasia was more common in the birds on the H feed than in the birds on the L feed. Hence, tibial dyschondroplasia was correlated with rapid growth. The other skeletal lesions did not differ in incidence in the birds on the H and L feeds. It was postulated that greater difference in growth rate is necessary for a demonstration of a possible correlation between skeletal deformities and growth rate.


Pathology of so-called leg weakness in the pig. The morphologic background of the poorly defined clinical entity called leg weakness in pigs was determined. Two hundred and thirty skeletons of breeding pigs slaughtered because of leg weakness were studied. Macroscopic, radiologic, bacteriologic and histologic examinations were performed. Three main types of pathologic changes were found and defined: Osteochondrosis, osteoarthrosis and arthritis. Distribution of the changes and their interrelationship were studied as well as age, sex and breed incidence. It was concluded that osteochondrosis is the most important cause of leg weakness. In adolescent animals, manifestations of osteochondrosis were seen in joints and growth plates of almost all individuals. A high frequency of osteoarthrosis, higher with increasing age, was also found. In most animals osteoarthrosis seemed to be secondary to osteochondrosis. However, primary osteoarthrosis of the hock joint similar to spavin in the horse was found. This lesion occurred already in young animals. Chronic proliferative arthritis and discospondylitis were frequent. Suppurative arthritis and osteomyelitis due to pyemia were also common findings. Inflammatory changes were seen as complications to osteochondrosis. Arthritis was secondary to osteochondrosis in importance as the cause of leg weakness in the pig.


Normal and pathologic skeletal development in broiler and leghorn chickens. A comparative investigation. Skeletal development of broilers of two lines of the Hybro Compact strain was compared to the skeletal development of leghorns. Environment, management, and feeds were identical for comparable groups of broilers and leghorns. Half of the broilers (representing both lines) and half of the leghorns were given a high energy feed, while the other half of the birds of both breeds were given a low energy feed. No great difference in feed conversion between broilers and leghorns was found. The broilers consumed more feed and increased their body weight and volume of the tibia almost twice as fast as the leghorns. The difference in longitudinal growth of the tibia of the birds of the two breeds was much less evident. Locomotor disturbances (leg weakness) were common in the broilers, and in the majority of the cases the problems were caused by skeletal deformities. The leghorns were almost completely free of skeletal deformities and had no locomotor disturbances. Likewise, dyschondroplasia was not found in the leghorns, while it was common in the broilers. Most of the skeletal deformities in the broilers were explained as the result of mechanic-traumatic factors, acting on a rapidly remodeling skeleton, apparently of insufficient strength for the heavy body weight. Dyschondroplasia, on the other hand, was considered to be caused by a generalized factor governing growth. Basic histologic differences between broilers and leghorns already at an age of 7 days supported this theory. It was concluded that both leg deformities and dyschondroplasia in the broiler are man made problems, caused by genetic selection for characteristics of economic importance, without enough consideration to function and health of the animals.


Morphology of osteochondrosis and sequelae in pigs. The morphologic features of osteochondrosis were studied in skeletons of 176 pigs of different ages. Macroscopic and histologic as well as radiologic and microradiographic methods were used. Osteochondrosis was found to be a generalized disease, characterized by a disturbance of endochondral ossification. It was concluded that early changes occur at about 4 months of age in the growth cartilage. Both the metaphyseal growth cartilage (growth plates) and epiphyseal growth cartilage (joint cartilage) were affected. Abnormal hypertrophy and hyperplasia of cartilage as well as focal degeneration were seen in the early cases. Osteochondrosis was found to lead to abnormal skeletal growth and change in shape of various bones and joints. Serious manifestations of osteochondrosis were epiphyseolysis and osteochondritis dissecans. A quantitative difference was found between different individuals in development of osteochondrosis. The onset of clinical signs was thought to depend on degree and localization of osteochondrosis.


Effects of vitamin D and A, calcium, phosphorus, and protein on frequency and severity of osteochondrosis in pigs. The generalized nature of osteochondrosis indicates that systemic factors, i.e. of nutritional or genetic origin, are of importance in etiology of osteochondrosis. Several experimental investigations are reported. On the basis of the morphologic findings it was considered possible that dietary factors such as hypovitaminosis D, hyper- or hypovitaminosis A, and low calcium or high protein intake could be of importance in etiology. The reason for this was that there were similarities between cartilage changes in osteochondrosis and those observed in various dietary imbalances in other domestic animals and in experiments with pigs. The effect of hyper- and hypovitaminosis A and hypovitaminosis D was studied in 23 pigs from 2 litters. In another experiment 32 pigs from 4 litters were fed rations with varying levels of calcium, phosphorus, and protein in the feed. Clinical examinations were made of these animals. Bone seeking fluorochromes were given prior to slaughter, and complete necropsy was performed, including macroscopic, radiologic, histologic and microradiographic techniques. Neither hypervitaminosis A, nor hypovitaminosis A and D had any noticeable effect on frequency and severity of osteochondrosis. A similar negative finding was made in the experiment with different intake of calcium and protein.


Study of spin-lattice and spin-spin relaxation times of 1H, 2H, and 17O in muscular water. Spin-lattice (T1) and spin-spin (T2) relaxation times of proton, deuteron, and oxygen-17 in muscle water have been measured at 9.21 MHz in the temperature range of 0 degree--40 degrees C. The values of the apparent activation energy for the three nuclei are (in kJ . mol-1) 9.1, 19, and 18 for 1/T1, and -1.3, 4.2, and 14 for 1/T2, respectively. The relatively small values for T2 for 1H and 2H and their low apparent activation energies are attributed to hydrogen exchange between water and proteins; this exchange does not affect the 17O relaxation. Quantitative calculations on deuteron T1 and oxygen-17 T1 and T2 have been made. The effect of surface-induced anisotropy on a minor fraction of water molecules is considered in some detail, and a new expression for its spectral density similar to that of liquid crystalline systems is applied in the calculation. It is suggested that water on the surfaces of macromolecules has a rotational correlation time of tau c approximately 1 x 10(-9) S, with a time constant of tau x approximately 3 x 10(-7) S, which is characteristic of the relaxation of the local structure.


Studies in epidemiology of maedi/visna in sheep. Lambs born to ewes from flocks with a high incidence of maedi/visna were separated from their dams at birth (group 1), or after 10 h (group 2), six weeks (group 3) or one year (group 4) and were observed for periods of up to eight years. Group 1 lambs remained free of infection while 28 per cent, 76 per cent and 81 per cent respectively of lambs in the other groups developed clinical, serological or histopathological evidence of infection during the observation period. It is therefore concluded that vertical transmission, if it occurs at all, is of little significance in the epidemiology of the disease. The number of serologically, virologically and histopathologically maedi/visna positive sheep, the time of onset of disease and the severity of lesions were related to the duration of exposure to the parent flock. In a separate trial no evidence was obtained for the transmission of maedi/visna by Muellerius capillaris larvae.


Calcium-dependent aldosterone secretion in anephric and nonnephrectomized patients on regular hemodialysis. The present study was undertaken to investigate the effect of a continuous calcium infusion on the plasma levels of aldosterone, renin activity, and cortisol in six anephric and four nonnephrectomized patients on regular hemodialysis. In both groups, a significant increase in whole blood ionized calcium (b-Ca2+) was demonstrated. A significant increase in plasma aldosterone (PAC) was noted in the nonnephrectomized patients, in whom the rise in PAC correlated with the increase in b-Ca2+. However, in the anephric patients only a smaller and insignificant increase in PAC was found. No significant changes were demonstrated in plasma cortisol or renin activity, nor in potassium or sodium concentrations in either group. It is concluded that ionized calcium influences the plasma levels of aldosterone in uremic patients on regular hemodialysis.


Thyrotropin displacement activity of serum immunoglobulins from patients with Graves' disease. A radioreceptor assay for human thyroid stimulators has been employed in various groups of patients. The ability of Ig to displace the labeled TSH from the receptors is referred to as "TSH displacement activity (TDA)." In active Graves' disease, TDA was above normal in 76% of the cases, and in the remaining patients, it was above the 76th percentile, suggesting that thyroid-stimulating Ig (TSIg) may have been present in all cases, but not always demonstrable by this method. Significant TDA was not found in normal persons or in toxic or nontoxic nodular goiters. It was also negative in some patients with "euthyroid ophthalmic Graves' disease." In patients with Graves' disease controlled with antithyroid drugs, positive TDA accurately predicted the recurrence of hyperthyroidism in eight of nine cases from whom the drugs were withdrawn. Thus, TSIg appears to be a prerequisite of the hyperthyroidism of Graves' disease. Moreover, the remission of hyperthyroidism was due to the disappearance of TSIg (immunological remission) in most cases in this small series. Serum TDA may provide a means of detecting immunological remission. The exophthalmos of Graves' disease does not require thyroid-stimulating Ig.


An increase of plasma triiodothyronine and thyroxine after administration of dexamethasone to hypothyroid patients with Hashimoto's thyroiditis. In an attempt to study the effect of adrenal steroids on plasma thyroid hormone concentration in patients with Hashimoto's thyroiditis, 2 mg dexamethasone was administered for 2-4 weeks to 9 patients with normal plasma TSH, 3 patients with compensatory increase of plasma TSH, and 10 patients with a marked elevation of plasma TSH. Dexamethasone depressed plasma T4, T3, and TSH and reduced thyroid size in nine euthyroid patients, whereas a similar treatment did not affect plasma T3 and T4, but reduced plasma TSH and thyroid size in three patients with compensatory increase of plasma TSH. In contrast, dexamethasone elevated plasma T3 and T4 in 10 hypothyroid patients with a marked elevation of plasma TSH. The increase of T3 was more than that of T4, suggesting preferential secretion of T3. Plasma TSH was reduced, but was still above normal after administration of dexamethasone. It is suggested that the intrathyroidal autoimmune processes inhibiting thyroid hormone synthesis are significant in patients with more severe Hashimoto's thyroiditis, and that dexamethasone stimulated hormone synthesis by depressing the autoimmune processes.


Measurement of plasma adenosine 3',5'-monophosphate. Currently used methods for plasma cAMP measurements are either tedious (chromatographic preparation of sample) or potentially inaccurate (direct assay of plasma samples). A rapid, simple, and accurate competitive binding assay for plasma cAMP, which does not require chromatographic preparation of the sample, has been developed. This procedure prevents destruction of plasma cAMP by utilizing both theophylline and EDTA in the collection of the blood sample. Human plasma contains variable amounts of cAMP-binding activity which interfere with the measurement of cAMP by the standard competitive binding assay. Our assay procedure removes this binding activity by precipitation of plasma proteins with perchloric acid. The normal fasting value (+/- SD) of plasma cAMP using this technique is 17.6 +/- 4.3 pmol/ml, which is identical to values obtained by methods utilizing chromatographic purification of samples (18.3 +/- 3.0). The fasting plasma cAMP of patients with hyperparathyroidism is normal (16.2 +/- 3.4), but patients with maturity-onset diabetes mellitus have fasting values significantly below normal (12.3 +/- 2.4).


Immunoreactive endorphins, lipotropins and corticotropins in a human nonpituitary tumor: evidence for a common precursor. The immunoreactive (RIA), ACTH, LPH, alpha endorphin (alpha End) and beta End in an extract of a human pancreatic islet cell carcinoma causing the ectopic ACTH syndrome were assayed after gel exclusion chromatography. In addition to "big", "intermediate" and "little" ACTHs, beta lipotropin (beta LPH) and gamma LPH, the eluate fractions contained RIA-alpha End and -beta End. The major RIA-beta End component appeared to be h beta LPH, which has beta End as its carboxy-terminus, but significant concentrations of a component that coeluted with synthetic p beta End were also found. Small amounts of RIA-alpha End were found in two peaks, one that may have represented 1-76 h beta LPH, with alpha End as its carboxy-terminus, and one probably representing alpha End itself. RIA-ACTH, -LPH and -beta End were found in the void volume fractions. Thus, a human nonpituitary tumor that produced ACTH and LPHs also produced endorphins, all perhaps deriving from a common precursor.


Proposed metabolic dysfunctions in diabetic microthromboses and microangiopathy. This report describes, at least in part, the role of prostaglandin and cyclic nucleotide metabolism in the etiology of the vascular disease associated with diabetes mellitus. Alterations in this metabolism seem associated with induction of platelet aggregation leads to microthromboses leads to microangiopathy sequences that are subtle but inexorable over a long period of time. Prostaglandins are generally elevated in blood from patients having frank signs of diabetic retinopathy when compared with nondiabetic subjects. Prostaglandin concentration remained elevated in diabetic retinopathy patients receiving indomethacin. We formed, therefore, the working hypothesis--yet to be fully tested either in patients or animal models with and without indomethacin treatment--that the increased prostacyclin (synthesized by endothelial microsomes) and cyclic-AMP production, both of which favor prevention of platelet aggregation, accompany the increased concentration of one or more of the prostaglandin E and F compounds. Concurrently, there may be an accompanying reduction of thromboxane A2 (synthesized by platelet microsomes) and cyclic-GMP (both of which favor platelet aggregation) production in the diabetic patients. The elevated prostaglandin in the diabetic patients not receiving indomethacin could possibly be directed toward slowing but not preventing the progression of the complex disease process in diabetes.


Serum angiotensin--converting enzyme (SACE) in sarcoidosis and other granulomatous disorders. Serum angiotensin-converting enzyme (SACE) activity was significantly higher in 90 patients with sarcoidosis (55 +/- [S.D.] 23 nmol min-1 ml-1) than in 80 healthy controls (34 +/- 9 nmol min-1 ml-1). Steroid therapy modified SACE activity; 60 sarcoidosis patients who were not being treated with steroids had significantly higher enzyme activities (58 +/- 24 nmol min-1 ml-1) than 30 steroid-treated sarcoidosis patients (40 +/- 19 nmol min-1 ml-1). In 50% of the non-steroid treated sarcoidosis patients SACE activity was more than than 2 S.D. above the mean value for the controls. SACE activity was measured in 22 tuberculous patients (38 +/- 14 nmol min-1 ml-1), 20 leprosy patients (34 +/- 9 nmol min-1 ml-1), 31 with primary biliary cirrhosis (44 +/- 20 nmol min-1 ml-1), 26 with inflammatory bowel disease (31 +/- 9 nmol min-1 ml-1), eight with hepatic granulomatous disease, five with Hodgkin's disease, and two with schistosomiasis. The combined false-positive rate for these non-sarcoidosis patients was 10%. Serial SACE assays provide useful information on the course of sarcoidosis and response to steroid treatment.


EEG sleep in primary depression. A longitudinal placebo study. Characteristic EEG sleep changes in depression are highlighted by a sleep continuity disturbance, delta sleep reduction, and a shortened REM latency. Since these findings have been derived primarily from only a few baseline recordings, questions regarding their persistence and/or variability have not been previously addressed. As part of an extensive set of investigations of EEG sleep in depression, we examined nightly the sleep of 12 hospitalized, non-delusional, primary depressives who were involved in a program of active psychosocial treatment intervention and received only placebo during a 5-week study period. EEG sleep findings revealed a relative lack of change across time, particularly in those parameters reported to be associated with a primary or 'biologic' depressive episode. While some degree of clinical improvement was noted, the group failed to achieve a state of remission or even partial remission as determined by the Hamilton Rating Scale. It appears that the major sleep alterations associated with such disorders persist for up to at least 5 weeks in the absence of pharmacologic or other somatic intervention.


Cystosarcoma phyllodes: a clinicopathological analysis of 14 cases. Experience with 14 cases of cystosarcoma phyllodes encountered at the 2nd Department of Propedeutic Surgery, Athens, University, between October, 1928 and December, 1978 is reported. Local excision including a wide cuff of uninvolved breast tissue is indicated for histologically proven benign and small tumors. For bulky or malignant tumors, a simple mastectomy is indicated. none of these neoplasms had metastasized to the axillary lymph nodes and, for this reason, and axillary node dissection was contraindicated.


Effect of oral glucose ingestion on renal phosphate reabsorption and clearance in vitamin D-resistant rickets. The effect or oral glucose ingestion on renal phosphate reabsorption was studied in 13 patients with the inherited form of vitamin D-resistant rickets (VDRR) and 5 normal subjects. In contrast to the normal subjects, glycosuria developed in six VDRR patients after glucose ingestion and resulted in a further 43% decrease in renal phosphate reabsorption. This was accompanied by a 33% increase in phosphate clearance. This was not attended by differences in fasting glucose or phosphorus levels between groups, or in their respective values 1 h after glucose ingestion. Baseline renal phosphate reabsorption was less and baseline phosphate clearance was greater in those VDRR subjects who developed glycosuria. The accumulated data suggest that excessive glucose ingestion by some patients with VDRR may add an additional insult to the phosphaturia characteristic of this disorder. This, in turn, would further compromise the response of circulating phosphate to therapeutic attempts at oral phosphate supplementation, thereby reducing the efficacy of oral phosphate therapy on skeletal growth and development in this disorder.


Effect of parathyroid hormone on plasma prolactin in man. The iv infusion of parathyroid extract or the synthetic fragments of 1-34 bovine or human parathyroid hormone produced a rapid and marked increase of plasma PRL in normal subjects. The stimulation of the release of endogenous parathyroid hormone by administration of disodium EDTA also resulted in a parallel increase of plasma PRL. Parathyroid hormone did not act via plasma cAMP, as the plasma level reached by this nucleotide was too small to produce PRL release. The ingestion of L-dopa 2 h before parathyroid hormone infusion suppressed the PRL response, suggesting that dopamine and parathyroid hormone interact at a common site. As it has been recently shown that PRL stimulates the renal synthesis of 1,25-dihydroxycholecalciferol, the present data suggest that the effect of parathyroid hormone on this synthesis may be due to the increase in plasma PRL.


Comparison of the corticotropic action of two synthetic, substituted analogues of ACTH: ACTH1--17 and ACTH1--18. The effect of a recently synthesized analogue of ACTH, Ala1, lys17-ACTH(1-17-4-amino-n-butylamide; (ACTH1-17) upon serum cortisol levels in 9 healthy males was compared with that of ACTH1-18, D-ser1-lys17, 18-ACTH(1-18). Both compounds induced an identical rise in serum cortisol which lasted for at least 8 hours. It is concluded that the amino acid in position 18 is not essential for the corticotropic properties of the synthetic heptadecapeptide. The prolonged effect of these compounds indicates a potential therapeutic significance for ACTH analogues of low molecular weight.


Regulation of the synthesis of human chorionic gonadotrophin by 5-bromo-2'-deoxyuridine and dibutyryl cyclic AMP in trophoblastic and nontrophoblastic tumor cells. Synthesis of hCG and its alpha subunit (hCG-alpha) by trophoblastic tumor cell lines (JEG-3, Reid, and BeWo) and by nontrophoblastic tumor cell lines [ChaGo and three HeLa lines (HeLa CCL2, HeLa S3, and HeLa 65)] was studied. cAMP, dibutyryl cAMP, and 8-bromo-cAMP induced, but 5-bromo-2'-deoxyuridine inhibited, the synthesis of hCG-alpha and hCG in trophoblastic tumor cells, Addition of 5-bromo-2'-deoxyuridine partially blocked the induction by dibutyryl cAMP in these cells. The synthesis of hCG-alpha and hCG in nontrophoblastic tumor cells was stimulated by dibutyryl cAMP. cAMP and 8-bromo-cAMP, however, did not affect the synthesis of hCG-alpha, although the synthesis of hCG was enhanced. 5-Bromo-2'-deoxyuridine induced hCG-alpha synthesis but did not affect hCG synthesis in nontrophoblastic tumor cells. In HeLa S3 and HeLa 65 cells, the amount of hCG-alpha and hCG produced in the presence of dibutyryl cAMP and 5-bromo-2'-deoxyuridine together was greater than the total hormone produced by each inducer separately. Our data indicate that the regulation of the production of hCG-alpha and hCG differs in trophoblastic and nontrophoblastic tumors.


Isolated deficiency of adrenocorticotropin (ACTH) and lipotropins (LPHs). A patient with hypoadrenocorticism was found to have low basal plasma concentrations of ACTH and lipotropins and deficient responses of these hormones to insulin-induced hypoglycemia and lysine vasopressin. The adequacy of secretion of other anterior pituitary hormones was assessed either directly, by measuring their concentration in plasma, or indirectly, by assessing end organ function, under basal and stimulated conditions. The responses of gonadotropins to LRH and of PRL and TSH to TRH were normal. The etiology of this rare condition of isolated deficiency of ACTH and lipotropins remains to be elucidated.


Reliability of the 30-minute ACTH test in assessing hypothalamic-pituitary-adrenal function. An insulin hypoglycemia test and a 30-min ACTH test were performed in 90 patients with proved or suspected hypothalamic-pituitary-adrenal hypofunction and in 10 normal subjects. The peak plasma cortisol concentration during hypoglycemia and the cortisol concentration at 30 min after injection of 250 micrograms corticotropin 1-24 were compared. A very close correlation (r = 0.92) was found between the the two concentrations, with no major discrepancy in any case. It is concluded that the short ACTH test accurately reflects the integrated hypothalamic-pituitary-adrenal function, as assessed by the insulin hypoglycemia test.


ACTH-secreting medullary carcinoma of the thyroid presenting a severe idiopathic osteoporosis and senile purpura: report of a case and review of the literature. A 64-yr-old female presented with severe osteoporosis and easy bruisability of over 2-yr duration. Biopsy of a neck mass revealed medullary carcinoma of the thyroid. Subsequently, lymphangitic pulmonary metastases were demonstrated which had been present radiographically for at least 4 yr. Basal serum calcitonin was markedly elevated and increased during calcium infusion. The diagnosis of ectopic ACTH syndrome was first entertained when hypokalemic alkalosis was observed during evaluation of her carcinoma. Elevated urinary 17-hydroxycorticosteroids, 17-ketosteroids, plasma cortisol, and immunoreactive plasma ACTH levels were documented. Adrenal steroidogenesis seemed to suppress on high dose dexamethasone. The primary tumor and its metastases contained high concentrations of immunoreactive ACTH and beta-melanocyte-stimulating hormone. Hepatic metastases contained extremely high concentrations of calcitonin. In contrast to the usual presentation of the ectopic ACTH syndrome as primarily hypokalemic alkalosis and glucose intolerance, patients with relatively benign and indolent ACTH-secreting tumors, such as certain cases of medullary carcinoma of the thyroid, may present with more typical signs and symptoms of Cushing's syndrome. The more pronounced cushingoid features in this latter group presumably reflects a more prolonged period of exposure to elevated glucocorticoid levels. Ten cases of ACTH-secreting medullary carcinoma of the thyroid from the literature are discussed. Extopic ACTH production by such tumors should be considered in the evaluation of patients with Cushing's syndrome or unexplained severe osteopenia.


A virilizing adrenocortical tumor in a female infant: in vivo and in vitro biochemical characteristics. Normal adrenal and adrenal tumor cells from a female infant with a virilizing adrenal tumor were grown in tissue culture as monolayers for a period of 7 weeks. Half of the cultures were exposed to ACTH (0.1 U/ml). The cells grew well and continued to produce steroid hormones over the entire period in culture. Production of steroid hormones was measured by RIA of individual steroids in the culture medium. Unstimulated normal and tumor cells produced equivalent amounts of cortisol, 11 beta, 18, 21-trihydroxy-4-pregnene-3,20-dione, and 18,21-dihydroxy-4-pregnene-3,20-dione, but tumor cells produced lesser amounts of dehydroepiandrosterone (DHA), androstenedione, testosterone, and progesterone. Normal cells exposed to ACTH showed an increase in all steroids measured, whereas ACTH-exposed tumor cells showed an increase principally in DHA consistent with a deficiency in 3 beta-hydroxysteroid dehydrogenase activity. Circulating levels of DHA, androstenedione, and testosterone were elevated in the patient before removal of the adrenal tumor. The production of androgens by tumor cells in vitro resembled the pattern of circulating steroids in vivo. These studies demonstrate that tissue culture of human adrenal cells provides a means both to determine their biochemical characteristics and to investigate their responses to exogenous hormones.


Relationship between thyroid status and Graves' disease-specific immunoglobulins. The prevalence of TSI in Graves' disease was investigated with a radioligand receptor assay using human thyroid membranes and highly purified labeled porcine TSH or bovine TSH in 110 patients before treatment and in 39 patients after successful treatment with antithyroid drugs. In 11 patients, the assay was performed before, during, and after therapy. The results in the radioligand receptor assay were compared to thyroid function tests (TRH test, free T4 index, serum %3 level, and thyroid suppression test). Normal IgG inhibits nonspecifically, but dose dependently, the binding of [125I]TSH to thyroid membranes. IgG samples from patients were only considered to be positive if they reduced the binding of the labeled hormone below the mean value minus 2 SD of the controls. TSI activity was found in 50% of the patients with untreated diffuse toxic goiter, in 3 out of 27 cases who regained suppressible thyroid function, and in 5 out of 12 cases who were off therapy for more than 6 weeks and showed normal TRH tests and normal hormone levels but in whom the suppression test was not performed. Our data cannot prove the hypothesis that only humoral antibodies are responsible for the thyroid stimulation in Graves' disease, but of course they cannot exclude it; furthermore, they suggest the existence of antibodies which bind to the human thyroid cell membrane without necessarily stimulating thyroid cellular activity.


Licorice raises urinary cortisol in man. The finding that urine cortisol excretion was increased in patients with hypokalaemic hypertension induced by licorice addiction led to this study on the effect of licorice in normal subjects. Thirteen normal volunteers ate either 100 or 200 g licorice for 1-4 weeks and assessment of pituitary-adrenal function was made before, during, and 1 week after cessation of licorice ingestion. Urine cortisol excretion more than doubled in 10 of 13 subjects (mean, 33.8 +/- 15.6 SD before and 83.3 +/- 56 SD micrograms/24 h at 1 week after commencing licorice) and excretion rates similar to those observed in Cushing's syndrome were seen in 7 subjects (range, 91-226, compared to normal range of 11-82 micrograms/24 h). Urine cortisol excretion remained significantly elevated (P less than 0.01) above control levels for at least 1 week after licorice was withdrawn. Despite these increases, urinary steroid metabolite (tetrahydrocortisol, tetrahydrocortisone, tetrahydro-11-deoxycortisol, 17-ketogenicsteroids, and 17-ketosteroids) excretion was not affected, plasma cortisol and ACTH values were unchanged, and normal 0800-1600-h diurnal variation of plasma cortisol was maintained. The direct intraadrenal infusion of the active mineralocorticoid component of licorice, glycyrrhetinic acid, in two sheep with autotransplanted adrenal glands failed to stimulate cortisol secretion acutely. It is concluded from these studies that the licorice-induced changes in cortisol excretion are not a result of adrenocoritcal stimulation but more likely represent a change in the renal handling of cortisol.


Role of angiotensin II in the aldosterone secretory response to adrenocorticotropin in sodium-restricted normal human subjects. A study was performed to determine the possible role of angiotensin II (AII) in mediating the increased adrenal aldosterone response to infused alpha 1-24-ACTH, induced by sodium deprivation. Nine normal subjects, aged 18-31 yr. received 8-h infusions of 1) alpha 1-24-ACTH (0.5 U given over 8 h) while on a diet with unrestricted sodium content; 2) saralasin at 0.5 micrograms/kg/min or ACTH alone (0.5 U over 8 h) on the 7th day of a 10 meq sodium diet; and 3) ACTH and saralasin together on the 8th day while still on sodium restriction. AII was administered for the last 2 h of infusion 3. Plasma cortisol and aldosterone concentrations were measured at hourly intervals during the infusions and plasma renin activity was measured at 2 hourly intervals. ACTH infusion produced an increase in the plasma aldosterone concentration which was significantly greater during sodium restriction than when sodium intake was unlimited. This increase was not associated with an ACTH-induced rise in the plasma renin activity and was not significantly altered when ACTH was administered with saralasin. The rise in plasma cortisol concentration induced by ACTH was not significantly different when the normal subjects were on liberal and restricted sodium intakes. It is concluded that AII plays little if any acute role in increasing the stimulatory action of ACTH on aldosterone secretion during sodium restriction.


Altered responsiveness to thyrotropin in thyroid slices of Graves' disease preoperatively treated with excess iodide. In a previous paper, we demonstrated that the acute administration of excess iodide inhibits the adenylate cyclase-cAMP system in mouse thyroid lobes. In the present study, we examined whether presurgical therapy with stable iodide reduces the responsiveness to TSH in thyroid tissues from patients with Graves' disease. Eight patients with Graves' disease were presurgically treated with methimazole and stable iodide and six were given methimazole alone. Normal tissues from five patients with thyroid nodules were also tested. We have found that stimulation by TSH (5 and 50 mU/ml) of cAMP formation in thyroid slices from patients preoperatively treated with methimazole and iodide is significantly less than in slices from patients treated with methimazole alone. Similar observations were also made with other thyroid stimulators, such as prostaglandin E2 and 4-methylhistamine. Furthermore, thyroid slices from patients treated with methimazole alone responded to TSH to the same degree as slices of normal tissues. The data suggest that one of the reasons for the hyporesponsiveness to TSH in thyroids from patients with Graves' disease is preoperative treatment with stable iodide.


Pituitary-adrenal axis activity in treated congenital adrenal hyperplasia: static and dynamic studies. The pituitary-adrenal axis activity was evaluated in 43 patients, treated for congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency, by measuring plasma ACTH, 17-hydroxyprogesterone (17-OHP), testosterone, and aldosterone. Dynamic studies were performed by injecting 250 micrograms synthetic ACTH im and collecting blood samples 1 h later for steroid analysis. Twelve to fourteen hours after the last hydrocortisone dose given the evening before, plasma ACTH fluctuated widely from less than 10-475 pg/ml, 17-OHP exceeded normal values and varied from 1-275 ng/ml, while testosterone ranged from 3-151 ng/100 ml. The correlations between ACTH and 17-OHP (n equal 61, r equal 0.665, P less than 0.001) and between 17-OHP and testosterone (n = 43, r = 0.761, P less than 0.001) were good, while that between 17-OHP and aldosterone (n = 64, r = 0.512, P less than 0.001) was rather poor. One hour after ACTH injection, the mean level of 17-OHP was significantly increased as compared to the mean basal level [96.8 ng/ml +/- 10.6 (SE) as compared to 67.0 ng/ml +/- 8.1 (SE)]. However, only 12 out of the 48 tests showed a positive response equal to or greater than 100%, and the majority of these responses (10 out of 12) occurred when basal levels of 17-OHP were between 10-70 ng/ml. This suggests that when basal levels fall outside these values, the pituitary-adrenal axis is either too inhibited or too stimulated to react to exogenous ACTH. Of the 48 tests where 17-OHP was measured, 23 had basal level values within these limits, the mean being 40.3 ng/ml. The corresponding mean ACTH level was 99 pg/ml with a wide range (1-230 pg/ml). On the other hand, in prepubertal children who exhibited 17-OHP concentrations between 10-70 ng/ml, testosterone varied from 3-30 ng/100 ml, with a mean of 16.0 ng/100 ml +/- 1.9 (SE) which is not different from the mean level found in normal children [14.0 ng/100 ml +/- 1.3 (SE)]. Thus, under the influence of endogenous ACTH which is moderately increased, 17-OHP concentrations far exceed normal values, whereas plasma testosterone seems to be unaffected.


Studies of the human testis. X. Properties of human chorionic gonadotropin receptor in adult testis and relation to intratesticular testosterone concentration. Receptors for [125I]hCG were found in adult human testis. The specific binding of [125I]hCG to testicular receptor is temperature dependent and is a saturable process with respect to added receptor protein and hormone. Scatchard analysis revealed a dissociation constant of 5.0 X 10(-10) M, and 6.2 fmol binding site/mg protein. Intact unlabeled hCG effectively inhibits the specific binding of [125I]hCG to human testicular receptors. For inhibition of binding of [125I]hCG, the alpha subunit has 3.0% of the potency of intact hCG and the beta subunit has 0.4% of the potency of intact hCG. Specific binding is pH dependent, with an optimum at pH 7.4. Brief exposure to extremes of pH causes irreversible damage to the receptors. Incubation with protease and trypsin results in an almost complete loss of binding activity, while ribonuclease, deoxyribonuclease, phospholipase C, or neuraminidase treatment does not significantly alter hormone-binding activity. Binding activity was found to be positively correlated to the concentration of intratesticular testosterone.