Search

The potential mechanisms whereby GABA mimetics and the antimanic agent lithium stabilize dopaminergic transmission are discussed. Evidence is presented that GABA mimetics, and in particular progabide, affect dopamine-mediated events in the basal ganglia on at least three levels. First, they reduce dopamine neuron activity in both the basal and the activated states. Secondly, on a long-term basis, they antagonize the proliferation of striatal dopamine receptors subsequent to chronic neuroleptic treatment. Thirdly, they modulate the expression of dopamine receptor activation by acting distally to the dopaminergic synapse. Lithium and GABA mimetics have the last two properties in common. These effects may represent the biochemical basis for the therapeutic action of GABA mimetics in iatrogenic dyskinesias. Moreover, the similarity between the biochemical effects of GABA mimetics and lithium suggest that the former drugs may have a therapeutic potential in mania.

Double-labeling experiments were performed at the electron microscopic level in the dorsal raphe nucleus of rat, in order to study the inter- and intracellular relationship of substance P with gamma-aminobutyric acid (GABA) and serotonin. Autoradiography for either [3H]serotonin or [3H]GABA was coupled, on the same tissue section, with peroxidase-antiperoxidase immunocytochemistry for substance P in colchicine-treated animals. Intercellular relationships were represented by synaptic contacts made by [3H]serotonin-labeled terminals on substance P-containing somata and dendrites, and by substance P-containing terminals on [3H]GABA-labeled cells. Intracellular relationships were suggested by the occurrence of the peptide within [3H]serotonin-containing and [3H]GABA-containing cell bodies and fibers. Doubly labeled varicosities of the two kinds were also observed in the supraependymal plexus adjacent to the dorsal raphe nucleus. The results demonstrated that, in addition to reciprocal synaptic interactions made by substance P with serotonin and GABA, the dorsal raphe nucleus is the site of intracellular relationships between the peptide and either the amine or the amino acid.

The binding of [3H]aldosterone in the chick intestine cytosol was analyzed in terms of affinity and specificity. In this tissue, aldosterone binds to the mineralocorticosteroid receptor, with a high affinity (Kd approximately 0.3 nM) and low capacity (approximately 50 fmol/mg protein), and to the glucocorticosteroid receptor. The selective labeling of the mineralocorticosteroid receptor was achieved by incubating the cytosol with [3H]aldosterone in the presence of RU 486. This synthetic steroid completely inhibited the binding of [3H]aldosterone to the glucocorticosteroid receptor and did not bind to the mineralocorticosteroid receptor. The oligomeric structure of the mineralocorticosteroid receptor was studied by using BF4, a monoclonal antibody which reacts with the 90-kDa heat shock protein (hsp 90), a nonhormone-binding component of nontransformed steroid receptors. The mineralocorticosteroid receptor sedimented at 8.5 +/- 0.4 S (n = 8) in a 15-40% glycerol gradient. This peak was shifted to 11.2 +/- 0.6 S (n = 5) after incubation with BF4, indicating that, in the cytosol, hsp 90 was associated with the mineralocorticosteroid receptor. Dissociation of the complex was observed on gradients containing 0.4 M KCl, as judged by the absence of displacement by BF4 of the 4.3 +/- 0.4 S (n = 10) peak. The effect of molybdate and tungstate ions, and of dimethyl pimelimidate, an irreversible cross-linking agent, on the stability of the hsp 90-receptor complex was investigated. Complexes recovered in the presence of 20 mM molybdate ions dissociated on gradients containing 0.4 M KCl (5.2 +/- 0.6 S (n = 4), whereas complexes prepared in the presence of 20 mM tungstate ions sedimented at 8.5 +/- 0.4 S (n = 7). Similarly, complexes prepared in the presence of molybdate ions dissociated during high pressure liquid chromatography (HPLC) gel filtration analysis performed in 0.4 M KCl (RS (Stokes radius) = 3.9 +/- 0.5 nm (n = 3) versus 7.3 +/- 0.2 nm (n = 3) in the presence of 20 mM molybdate ions), whereas complexes prepared in the presence of tungstate ions did not dissociate (RS = 6.9 +/- 0.2 nm (n = 3]. As observed for the tungstate-stabilized receptor, the cross-linked receptor dissociated neither on gradient containing 0.4 M KCl (9.5 +/- 0.1 S (n = 3] nor during HPLC performed in 0.4 M KCl (RS = 6.5 +/- 0.3 (n = 4]. Furthermore, the cross-linked receptor was more resistant to the inactivating effect of urea on aldosterone binding than the noncross-linked receptor prepared in the presence of either molybdate or tungstate ions.

Adrenalectomized rat kidney is commonly used for the study of mineralocorticoid mechanism of action in mammals. In this model, aldosterone is known to bind to two classes of binding sites: type I (mineralocorticoid) and type II (glucocorticoid). The study of the aldosterone binding in normal rat kidney requires the elimination of endogenous hormones bound to each type of receptor. Thus, a suitable technique was developed using in situ perfusion of the kidneys. The efficacy of this method was of about 85 to 90% at the level of both cytoplasm and nucleus. Aldosterone binding capacity was checked in normal rat kidney after in situ perfusion and was found to be 300 to 500% lower than in adrenalectomized rat kidney, both in cytoplasm and nuclei. Computer analysis of aldosterone binding parameters in the cytoplasm (30,000 X g supernatant) of rat kidney suggested that adrenalectomy might induce an important rise in the number of mineralocorticoid receptors (congruent to 260%). An increase in the number of glucocorticoid receptors was also observed but appeared to be lower. Aldosterone, when perfused during 24 h in adrenalectomized rats, lowered the number of type I sites to the same level as observed in normal rat kidney. This effect was fully reversible after interruption of aldosterone perfusion. These results suggested an aldosterone-induced down regulation of mineralocorticoid receptors.

Repeated treatment with haloperidol or lesion of nigrostriatal dopaminergic neurons with 6-hydroxydopamine produced a reduction in substance P immunoreactivity in the rat substantia nigra. This reduction was reversed by the repeated administration of progabide, a selective GABA receptor agonist. As GABA inhibits substance P release, these results suggest that the reduction in nigral substance P levels was due to an increased liberation of the peptide probably related to deficient GABAergic function induced by impairment of striatal dopaminergic transmission.

Metabolism of 9alpha-fluorocortisol (9alpha-F) has been studied in rat kidney slices, homogenate, and in the isolated perfused kidney. These studies show that the rate of 9alpha-F metabolism varies depending upon the experimental conditions. The major metabolite formed, identified by mass spectrometry, is 20(xi)-dihydro-9alpha-fluorocortisol. The kidney slice experiments show that only 3H-9alpha-F and none of the metabolites bind to cytosolic receptors. In competition experiments performed with tritiated and unlabeled 9alpha-fluorocortisol, aldosterone (A), dexamethasone (DM) and triamcinolone acetonide (TA), 9alpha-F was found to bind to mineralocorticoid receptors with a lower affinity than A but also to glucocorticoid receptors with a higher affinity than A. The Scatchard plot analysis indicated that 3H-9alpha-F is characterized by KD : 8.6 X 10(-9)M and N : 1.9 x 10(-13)moles/mg of protein. In conclusion it is felt that 9alpha-F would not be a better "marker" than aldosterone for the renal mineralocorticoid receptors.

The functional importance of hsp 90 interaction with steroid hormone receptors in hormone action has been previously proposed. This hypothesis, although not yet proven, is supported by new data obtained in our and other laboratories, whereas no conflicting experimental result has been presented. Our recent studies deal with results of hsp 90 cloning, normal and mutated receptor transfection experiments, effects of antihormone RU 486 and immunohistochemical data.

The specific early inhibition of DNA synthesis in reovirus-infected cells suggests that the cell nucleus is a target for virus-induced damage. We have now examined the affinity of reovirus proteins for DNA, postulating that such affinity could provide a mechanism for the inhibition. Cytoplasmic and nuclear extracts of cells labeled with [35S]methionine from 6 to 8.5 h after infection at high multiplicity were subjected to chromatography on denatured DNA – cellulose columns. Fractions from both cytoplasm and nucleus eluted with 0.6 M NaCl contained a protein with the same electrophoretic mobility on polyacrylamide slab gels as the nonstructural (NS) reovirus protein of the sigma size class. The protein also exhibited affinity for native DNA – cellulose and denatured DNA – agarose. Electrophoretic analysis in tube gels of cell extracts labeled for 48 h before infection with [14C]leucine and from 6 to 8.5 h after infection with [3H]leucine showed increased 3H label in this protein indicating it is reovirus specific. Small amounts of mu proteins also had DNA affinity. Purified virus did not bind strongly to DNA, suggesting that the binding protein is not a structural protein of the sigma size class on the outer surface of the virus. Our results provide evidence that the sigma NS protein binds to DNA. This affinity could interfere with chromosome function in the infected cell.

Substance P is a neuropeptide released in vivo from the substantia nigra, the principal substance P nerve terminal region in the rat brain. Its inactivation was investigated in a purified nigral synaptic membrane preparation. The membrane-bound enzyme shares many features with the endopeptidase 24-11 (EC 3.4.24.11): 1) hydrolysis of peptide bonds Gln6-Phe7, Phe7-Phe8 and Gly9-Leu10, 2) sensitivity to the inhibition by phosphoramidon and 3) relative affinity for substance P. Bestatine and captopril inhibit only the hydrolysis of the metabolites. These results suggest that substance P is inactivated in substantia nigra by endopeptidase 24-11 and that a bestatin-sensitive aminopeptidase and angiotensin converting enzyme may play a role in subsequent degradation of the substance P metabolites.

The potential mechanisms whereby GABA mimetics and the antimanic agent lithium stabilize dopaminergic transmission are discussed. Evidence is presented that GABA mimetics, and in particular progabide, affect dopamine-mediated events in the basal ganglia on at least three levels. First, they reduce dopamine neuron activity in both the basal and the activated states. Secondly, on a long-term basis, they antagonize the proliferation of striatal dopamine receptors subsequent to chronic neuroleptic treatment. Thirdly, they modulate the expression of dopamine receptor activation by acting distally to the dopaminergic synapse. Lithium and GABA mimetics have the last two properties in common. These effects may represent the biochemical basis for the therapeutic action of GABA mimetics in iatrogenic dyskinesias. Moreover, the similarity between the biochemical effects of GABA mimetics and lithium suggest that the former drugs may have a therapeutic potential in mania.

National audience

[ 3 H]-Aldosterone and [ 3 H]-dexamethasone binding characteristics in the kidney cytosol of adrenalectomized rats were assessed during normal growth and compensatory hypertrophy of the kidney. The nephrectomy was performed on one month old rats and the growth processes studied for four weeks. The models for the ligand interaction with the cytosolic receptors, the corresponding dissociation constants and the maximum number of sites for each class of receptor were all determined. The dissociation constants of aldosterone ( KD I ≈- 1.4 × 10 −9 mol/l, KD II ≅ 4.3 × 10 −8 mol/l) and dexamethasone ( KD ≅ 6.5 × 10 −9 mol/l) as well as the tissue concentrations of aldosterone binding sites (N 1 11 × 10 −13 mol/g tissue, N 2 ≅ 194 × 10 −13 mol/g tissue) and dexamethasone binding sites (N ≅ 265 × 10 −13 mol/g tissue) remained unchanged during normal growth and compensatory renal hypertrophy and were found identical to those previously determined for the adult adrenalectomized rat kidney. These results indicate a correlation between the total number of binding sites and renal growth whether the growth is normal or related to a hypertrophy.

Summary We have studied several aspects of the inhibition of DNA synthesis in Mengovirus-infected cells. In mouse L-929 cells at 5 h after infection at a multiplicity of 200 p.f.u./cell, there was a decrease in DNA synthesis. However, the rate of DNA replication fork movement, measured by equilibrium sedimentation of DNA sequentially labelled with BrdUrd and 3H-thymidine, was not significantly reduced in infected cells despite an 84% inhibition of DNA synthesis. Enzymic assay of the total cellular dTTP pool size showed it to be increased 37% in infected cells compared to controls. This was not accounted for by enhanced entry of thymidine nucleotides into the pool from either exogenous or endogenous sources, since the buoyant density of DNA pulse-labelled with 3H-BrdUrd was the same in infected and uninfected cells and there was no evidence for breakdown of DNA in infected cells. There was also a moderate decrease in the uptake of exogenous thymidine into the cell, shown by a reduction in the Vmax for transport with little change in the K m. By 6 h after infection, the dTTP pool size was slightly smaller in infected cells than in the controls and there was a marked inhibition in the rate of replication fork movement. These results show that Mengovirus infection blocks the entry of dTTP into DNA. At 5 h after infection, this block is manifested by an inhibition of initiation of new DNA chain synthesis. By 6 h, replication fork movement is also retarded and there is a more generalized derangement in DNA synthesis.

The numbers of type I and type II aldosterone receptors in the kidney cytosol of adrenalectomized rats were estimated after animals were treated with various steroids, or fed with high or low potassium diets. Oestradiol and 5 beta-pregnane-3,20 dione, which exhibited no affinity for aldosterone receptors, did not modify the levels of type I or type II receptors. Cortisol, corticosterone, progesterone and spirolactones, which all competed with aldosterone for both types of receptor, reduced the number of type I sites, as does aldosterone itself. Steroid treatment has no appreciable effect on type II receptors. We conclude that type I receptors are modulated by steroids able to bind to aldosterone receptors and that steroid-receptor interaction is an essential step in the receptor modulation process. The effects of potassium on aldosterone receptor modulation were tested in adrenalectomized rats on hypo- or hyperkalaemic diets. No change in receptor levels was observed in the rats on a low potassium diet, but the number of type I receptors increased in animals on a high potassium diet. However, the effects of potassium on receptor modulation were of lesser magnitude than those of aldosterone agonists and antagonists.

Publisher Summary This chapter discusses the functional similarities between benzamides and other neuroleptics. In contrast to classical neuroleptics, thioridazine, clozapine, and sulpiride fail to induce catalepsy in experimental animals and rarely cause extrapyramidal side-effects in man. It has been postulated that preferential presynaptic action of atypical neuroleptics may enhance dopamine release from nigrostriatal dopaminergic neurons and consequently, overcome the blockade of postsynaptic receptors responsible for extrapyramidal alterations. Among the various criteria for dopamine receptor classification, the one most widely used distinguishes between the receptors which are coupled to adenylate cyclase and those which are not. Accordingly, the former population is termed D 1 receptors while the latter are D 2 receptors. The chapter also discusses the effects of neuroleptics on some aspects of central noradrenergic transmission. The chapter presents as well the comparison of the effects of neuroleptics after their repeated administration. After repeated administration, the biochemical and behavioral effects of neuroleptics tend to attenuate as compared to a single treatment.

Specific binding of [3H]aldosterone to the cytosolic fraction of epithelial cells was studied in the human colon and terminal ileum. Analysis of [3H]aldosterone binding to the epithelial cells of ascending colon, caecum and ileum as a function of [3H]aldosterone concentration revealed only one class of specific receptors with an affinity constant of about 2 nmol/l. [3H]aldosterone binding was approximately the same in the sigmoid, descending and transverse colon and in the caecum, but slightly lower in the ascending colon and ileum. The specificity of the [3H]aldosterone binding was the same along the colon. The relative order of potency in inhibiting [3H]aldosterone binding was: aldosterone = SC 26304 = dexamethasone much greater than dihydrotestosterone greater than estradiol = RU 26988.

The short-term effects of chrysotile asbestos before and after SO2 sorption are studied in the rabbit after intratracheal injection of low doses of these pollutants. Chrysotile, as well as SO2-chrysotile, induces an increase in the unsaturated fatty acid content of lung surfactant, which is similar to that observed in the respiratory distress syndrome of the newborn, and an increase in the protein level of pulmonary washings which may be explained by an increase of the permeability of the blood-air barrier. These soluble proteins can interact with the surfacant and thus decrease its tensio-active properites.

Spirolactones are antimineralocorticoid compounds, which were introduced into therapeutics in 1960. They are useful in the treatment of hyperaldosteronism and essential hypertension, although their side effects (gynecomastia, impotence, menstrual cycle abnormalities) limit their use. They are known to antagonize the effect of aldosterone at the level of the target cells, and until recently this mechanism was the only one generally implicated. In 1972, Erbler showed that spironolactone inhibits aldosterone biosynthesis in vitro [1]. Since then, several authors have confirmed this effect, both in animals and man, and have therefore challenged the relative importance of peripheral aldosterone antagonism in favor of an inhibitory effect of these drugs on aldosterone biosynthesis. This editorial review attempts to clarify the findings and to determine whether antialdosterones act mainly at the target cell, or through inhibition of aldosterone production, or alternatively by a combination of each effect.

Repeated administration to rats of SCH 23390, a specific antagonist of the D-1 dopamine receptor, produced an increase in the substance P immunoreactivity in the striatum but not in the substantia nigra, whereas similar treatment with sulpiride, a specific D-2 dopamine receptor antagonist, reduced the nigral but not the striatal content of the peptide. When the two antagonists were given together, the SCH 23390-induced increase in striatal substance P was significantly reduced. The SCH 23390-induced increase in striatal substance P was curtailed by concomitant administration of progabide, a selective γ-aminobutyric acid (GABA) receptor agonist. These results suggest the existence in the nigro-striatal complex of two different substance P-containing neurons which are differentially regulated by the dopamine receptor subtypes and indicate a role of GABA in the action of SCH 23390.

The role of the soluble pool (cytoplasmic or cytosolic) of [3H]-aldosterone binding sites in the toad bladder was assessed by the use of two spirolactones, prorenone and spironolactone as a reference drug. Prorenone fulfills all the criteria for a specific competitive antagonist of aldosterone for its effect on Na+ transport. Compared with spironolactone (Ki approximately equal to 1 microM), prorenone was about eightfold less potent (Ki approximately equal to 8 microM). Competition for [3H]aldosterone binding sites by spironolactone and prorenone revealed an order of potency (spironolactone greater than prorenone) that corresponded to their antagonist activities in the Na+ transport assay. There was a linear correlation between the effects of the two spirolactones on the aldosterone-stimulated Na+ transport and their ability to displace [3H]aldosterone from its binding sites in the soluble pool. Finally [3H]prorenone binding sites were detected in the soluble pool but an insignificant number of antagonist-receptor complexes were found associated with the nuclear pool. Our study indicates that the aldosterone binding sites of the soluble pool are indeed mineralocorticoid receptors, which are probably the first intracellular mediators leading to an increased Na+ reabsorption.

The behavior of mineralocorticoid and glucocorticoid receptors of rabbit kidney cytosol was investigated on two affinity gels: a new affinity matrix prepared with a 3-O-derivative of carboxymethyloxime deoxycorticosterone (deoxycorticosterone gel) and a gel linked to a 17 beta-dexamethasone derivative (dexamethasone gel). Deoxycorticosterone gel was highly specific, since it retained mineralocorticoid but not glucocorticoid receptors, and dexamethasone gel exhibited high selectivity for glucocorticoid receptors since it did not bind mineralocorticoid receptors. The use of these two matrices allowed separation of mineralocorticoid and glucocorticoid receptors and further characterization of each type of cytosolic receptors after its isolation. Cytosolic mineralocorticoid and glucocorticoid receptors stabilized by tungstate were found to have a Stokes radius of approximately 6 nm, as determined by high performance size exclusion chromatography and a sedimentation coefficient of approximately 9 S, determined on a glycerol density gradient containing tungstate, under either high or low salt conditions. The hydrodynamic parameters, binding characteristics, and specificity of mineralocorticoid receptors were the same in the untreated and dexamethasone gel-treated cytosol. Similarly glucocorticoid receptor characteristics remained unchanged after deoxycorticosterone gel treatment, indicating biochemical independence of cytosolic mineralocorticoid and glucocorticoid receptors. The [3H]aldosterone receptor complex eluted from deoxycorticosterone gel was recovered with a 30-40% yield and a purification factor of about 1000. Purified mineralocorticoid receptors had the same sedimentation coefficient as cytosolic mineralocorticoid receptors (9 S) but a different Stokes radius (4 versus 6 nm). The decrease in the Stokes radius of the purified mineralocorticoid receptors was probably due to the gel filtration method. These results indicate that the newly synthesized matrix specific for mineralocorticoid receptors constitutes a powerful tool for their extensive purification.

The effects of prolonged treatment with dietary lithium and/or haloperidol (infused by means of osmotic minipumps) on biochemical parameters indicative of striatal dopamine target cell supersensitivity have been investigated in the rat. When given concomitantly with haloperidol, lithium failed to prevent the fall of striatal dopamine metabolites observed 2 days following withdrawal and the tolerance to the elevation of dopamine metabolites in response to challenge with the neuroleptic during withdrawal. Prolonged treatment with lithium also failed to modify the changes in nigral dihydroxyphenylacetic acid levels and in striatal acetylcholine levels which occur under chronic neuroleptic treatment. Chronic dietary lithium alone consistently elevated substance P levels in substantia nigra. The usual decrease in nigral levels of the peptide that occurs in response to chronic treatment with haloperidol was prevented in animals treated concomitantly with lithium. These data suggest that the mechanism whereby lithium stabilizes dopaminergic supersensitivity does not seem to involve an action of the compound on the neuronal mechanisms regulating the activity of the nigro-striatal dopaminergic system or on striatal cholinergic neurons but may be related to the restoration of normal striato-nigral substance P ergic transmission.

1. The presence of high-affinity sites for [3H]-aldosterone was shown in the normal human renal tissue. 2. [3H]Aldosterone and [3H]dexamethasone binding were studied in human renal adenocarcinoma and in uninvolved external cortex, in 22 patients undergoing nephrectomy for renal adenocarcinoma. Tissue incubations were performed with either [3H]aldosterone (5 × 10−10 mol/l; 5 × 10−9 mol/l in the presence of unlabelled glucocorticoids) or [3H]dexamethasone (5 × 10−9 mol/l). 3. Cytosol [3H]aldosterone binding was six- to seven-fold lower (P < 0·001) in neoplastic than normal tissue. [3H]Dexamethasone binding was about twofold higher in neoplastic than in normal tissue. This difference was not significant. 4. Nuclear uptake experiments showed that, both in cytosol fractions and nuclei, [3H]aldosterone binding was lower in adenocarcinoma than in normal cortex. 5. The very low binding of [3H] aldosterone suggests that mineralocorticoid receptors are absent in renal adenocarcinoma, an hypothesis in line with the proximal origin of these tumours.

Eight hundred and ninety-nine men were studied, aged 16-65 inclusive, who underwent an elective inguinal herniorrhaphy during 1970 and 1971 in eight hospitals in Wessex, and under nine consultant surgeons. There was a significant variation in postoperative inpatient stay and total sickness absence between hospitals and between consultants. The physical activity involved in the patient's occupation, his age at operation, previous sickness absence, bilateral herniorrhaphy, attendance at follow-up outpatients' clinic, type of repair, and the influence exerted by three hospitals and two consultants accounted for only 21% of the variation in total sickness absence. The general practitioners who had referred patients to the hospitals for herniorrhaphy, and the consultant surgeons who carried out the operations, were sent a questionnaire to ascertain their attitudes towards follow-up outpatient appointments and the various factors identified in the first part of the study as significantly influencing total sickness absence. A higher proportion of GPs who felt that an outpatient appointment was necessary before return to work was found in relation to the patients who had the longest mean total sickness absences than among the GPs who looked after the patients with shorter total sickness absences.

In the urinary bladder of the toad Bufo marinus triiodothyronine selectively inhibits the late effect of aldosterone on Na+ transport. We have investigated whether T3 might mediate its antimineralocorticoid action by controlling: i) the level of aldosterone binding sites in the soluble (cytosolic) pool isolated from tissues treated with T3 (60 nM) for up to 20 hr of incubation; ii) the kinetics of uptake of 3H-aldosterone into cytoplasmic and nuclear fractions after 2 or 20 hr of exposure to T3. The number and the affinity of Type I (high affinity, low capacity) and Type II (low affinity, high capacity) cytosolic binding sites (measured at 0 degrees C) did not vary significantly after 18 hr of exposure to T3, while aldosterone-dependent Na+ transport was significantly inhibited. In addition, T3 did not modify the kinetics of uptake (90 min) of 3H-aldosterone into cytoplasmic and nuclear fractions of toad bladder incubated in vitro at 25 degrees C. By contrast, aldosterone itself was able to down-regulate its cytosolic and nuclear binding sites after an 18-hr exposure to the steroid hormone (10 or 80 nM). T3 slightly (20%) but significantly potentiated the down regulation of nuclear binding sites. In conclusion, T3 does not appear to have major effects on the regulation of the aldosterone receptor, which could explain in a simple manner its antimineralocorticoid action.

An experimental study has been carried out using different biological and biochemical in vivo and in vitro tests for assessing the toxicity of natural UICC (A) chrysotile and SO2-sorbed UICC (A) chrysotile and for detecting a possible synergistic effect. For in vivo studies, rabbits received an intratracheal injection of chrysotile fibers suspended in physiological saline (PS). The control group received only PS; all animals were sacrificed after 68 hr. The alveolar free cells were harvested by pulmonary lavage. The results have shown that chrysotile induces a decrease in the free cell population but there was no significant difference in the number of viable cells or in the nature of cells harvested between the two chrysotile groups. Enzymatic activities of the alveolar macrophages from animals injected with SO2-sorbed chrysotile showed a significant increase of the enzymes LDH and acid phosphatases. The LDH increase could be related to the affinity of the enzyme regarding some chemical forms of SO2. In vitro studies using alveolar macrophages harvested by pulmonary lavage have shown no differences between the two chrysotile groups when cell viability and enzyme release were studied. The toxic effect was due to chrysotile fibers (decrease of 38% in cell viability and increase in cellular enzyme release when compared with control). When rabbit red blood cells were used, both natural and SO2 chrysotile fibers showed the same hemolytic activity. The failure to detect high differences between the two chrysotile groups may be related to the chemical form of sorbed SO2.

In rats allowed access to food, and in food-deprived rats, fenfluramine (20 and 100 mg kg-1) and amphetamine (10 and 20 mg kg-1) provoked a hypotriglyceridaemic effect. No changes in plasma cholesterol concentration were observed. The time course of the absorption of a lipid load differed according to the nutritional status of the animals; being bellshaped under fed, and curvilinear under fasted, conditions. However, absorption under both nutritional conditions was inhibited by amphetmine and fenfluramine. When rats which had received the test compounds were administered glycerol trioleate containing a tracer dose of glycerol [1-14C]-trioleate or [2-3H]-glycerol trioleate, there was an inhibition in the increase of plasma radioactivity only in the case when the fatty acid contained the radioactive label. The net effect of lipid absorption was a transfer of dietary lipid from the gut to adipose tissue stores. There was never more than 5 per cent of the administered load in the liver. These observations indicate that amphetamine and fenfluramine may have acute effects in reducing circulating triglycerides, separate from the effects on lipid absorption from the gut. In this latter, the palmitoyl-CoA monooleinacyltransferase enzyme probalby plays a key role and appears a major target of the overall anti-obesity of fenfluramine.

Synchronizing each frame of the cinema of a moving animal with the corresponding EMG of some muscles permits a precise interpretation of the variations of the electric potentials in that muscle with each phase of the movement. e.g. in the rabbit, three bundles of the masseter muscle function independently during the mastication; in the pigeon, electric potentials appear in both ilio-tibialis and biceps femoris muscles some frames before the actual raising of the leg, when the animal on its perch becomes out of balance.

The cineradiographic technique, whereby, at 64 frames/second, the successive positions of every bone segment of animals in motion can be accurately analyzed, was used to study the vertical leap of a Galago alleni from the ground position. This jump, 13 to 14 times the body length of the animal excluding the tail, is a record among primates challenged only by the tarsier. Such vertical leaps from the ground are a common mode of locomotion for the galago. Two different patterns were observed, a symmetrical and an asymmetrical jump. In the former, the two hind limbs move in the same manner and simultaneously; and the propulsive force is equally distributed in both limbs. In the latter (more frequent in our records), the animal shifts its weight onto one foot during the preparatory phase and lifts the other; thus, only one of the hind limbs is responsible for the launching. The angular variations of every knee and ankle articulation and the successive positions of the hind limb bones were measured and diagrammatically analyzed frame by frame. A short film was made to illustrate the cineradiographic technique; it cinematographically and cineradiographically records in successive sequence the galago's leap from the ground. The symmetrical and asymmetrical jumps are also schematically presented.

The levels of noradrenaline, neuropeptide Y, 5-hydroxytryptamine, and substance P were measured and compared between the large arteries of the circle of Willis and the small cerebral vessels of the pia mater in the rat, rabbit, cat, and monkey. In all species, noradrenaline and neuropeptide Y concentrations were greater in the larger arteries than in small pial vessels. Noradrenaline concentrations were dramatically reduced following cervical sympathectomy, with the extent of diminution differing greatly in the various species; the effects of cervical ganglionectomy on neuropeptide Y concentrations were less pronounced. 5-Hydroxytryptamine concentrations in rats, cats, and rabbits were significantly greater in the small pial vessels, although measurable concentrations existed in the circle of Willis. In cats and monkeys, substance P was found in major arteries, but was not detectable at the level of the small pial vessels. The differences in the regional distribution of the various neurotransmitter candidates in the cerebrovascular bed may reflect their physiological significance.

We have examined the distribution of histone H1 in oocytes and preimplantation embryos of the mouse, using a polyclonal antibody raised against the histone H1 sub-types present in somatic cells. Immunofluorescence and immunoblotting analyses failed to detect somatic histone H1 in germinal vesicle (GV)-stage oocytes. In contrast, somatic histone H1 was detectable by immunofluorescence in the nuclei of GV oocytes previously injected with histone H1 as well as the nuclei of ovarian granulosa cells, and by immunoblotting in 8-cell embryos. 1- and 2-cell embryos examined by immuofluorescence did not contain detectable somatic histone H1. At the early 4-cell stage (54–56 hours post-hCG), 5 of 52 embryos contained somatic histone H1 in one or more nuclei. By the late 4-cell stage (66–68 hours post-hCG), however, 58 of 62 embryos contained somatic histone H1. In 8-cell embryos, morulae and blastocysts, all nuclei contained somatic histone H1 in every case. When embryos were exposed to the transcriptional inhibitor, -amanitin, beginning at the late 2-cell stage, they cleaved to the 4-cell stage but fewer than 10% developed histone H1 immunoreactivity. When treatment began at the early 4-cell stage, the embryos that remained at the 4-cell stage in the presence of the drug developed histone H1 immunoreactivity in half of the cases. Embryos that reached the 5-to 8-cell stage in the presence of the drug developed histone H1 immunoreactivity in every case. The protein synthesis inhibitor, puromycin, prevented development of histone H1 immunoreactivity in most embryos when added either at the late 2-cell or early 4-cell stage. When embryos were exposed to the DNA replication inhibitor, aphidicolin, beginning at the late 2-cell stage, they cleaved to the 4-cell stage, but developed only a very weak histone H1 immunoreactivity. These results indicate that oocytes and 1- and 2-cell embryos contain little or no somatic histone H1, which may imply that these cells contain immunologically distinct histone H1 subtypes. The somatic subtypes first appear at the 4-cell stage, through a process requiring embryonic transcription and DNA replication during the third cell cycle. These results suggest that the deposition of somatic histone H1 on chromatin is developmentally regulated during mouse embryogenesis.