An intraperitoneal administration of thyrotropin-releasing hormone (TRH, 2 or 20 mg/kg) produced behavioral excitement and consequently an increase in ANIMEX counts 15 min after the injection in a dose-dependent manner in 70-day-old rats. On the other hand, TRH (2 or 20 mg/kg)-induced behavioral excitement appeared more slowly (90-150 min) and more persistently in 7-day-old animals than in 70-day-old animals. TRH (2 or 20 mg/kg) significantly reduced specific [³H] spiperone binding to striatal membranes in 7- and 70-day-old rats compared to control, when the binding was examined in the membranes obtained from animals sacrificed 15 min and 150 min following TRH injection. In vitro addition of TRH up to 0.1 mM did not affect [³H] spiperone binding on days 7 and 70. From these results, it is suggested that striatal dopamine receptors could be involved in TRH-induced behavioral excitement in developing rats.

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Treatment of day-4 chick embryos with 6-aminonicotinamide (6-AN) impairs limb chondrogenesis and produce micromelia. However, treatment of day-10 chick embryos with 6-AN does not produce micromelia. In the present study, the glycosaminoglycan (GAG) biosynthesis in the cartilage isolated from the day-10 chick embryos treated with 6-AN in vitro and in the cartilage from the day-11 chick embryos treated with 6-AN in ovo at day-10 was examined. In the epiphyseal cartilage treated in vitro, the radioactivity incorporated into the GAG fraction was significantly decreased and the ³⁵S/³H ration in the GAG fraction and the molecular size of GAG chain were also decreased. However, the percent distribution of ΔDi-OS was almost unchanged. In the epiphyseal and diaphyseal cartilage treated in ovo at day-10, the alteration in GAG biosynthesis was not observed except the reduction of the molecular size of GAG in epiphyseal cartilage. Furthermore, the biosynthetic activity of GAG in the process of recovery/repair was also examined using the micromelial cartilage isolated from day-11 chick embryos given 6-AN in ovo at day-4. In the diaphyseal cartilage, the radioactivity incorporated into the GAG fraction and the ³⁵S/³H ratio were increased, whereas the percent distribution of ΔDi-OS was almost unchanged. These results indicate that treatment of day-4 chick embryos with 6-AN produces the rebound phenomena in the diaphyseal cartilage isolated at day-11. In the present study, it has become clear that 6-AN directly impairs GAG biosynthesis in epiphyseal cartilage and that the effects of 6-AN on isolated cartilage differ from the effects of 6-AN on limb mesoderm in ovo. Thus, the defects induced by 6-AN in isolated cartilage cannot be conceived of as the cause of the bone malformation asserted by Seegmiller et al.

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The effect of cyclic GMP on the accumulation of p-aminohippurate (PAH) was investigated by using rat kidney corrical slices. When the slices were preincubated with cyclic GMP and thereafter incubated in the absence of the cyclic nucleotide for the assay of PAH transport, cyclic GMP inhibited PAH accumulation in the slices in a dose-dependent fashion. Pretreatment of the slices with dibutyryl cyclic GMP induced no inhibition of PAH uptake, but that with the combination of cyclic GMP and butyrate inhibited the accumulation to the same extent as did cyclic GMP. Guanosine and its monophsphate (GMP) had no effect on PAH accumulation in the slices. Inhibitory effect of cyclic GMP on PAH accumulation was not due to the changes of electrolyte content and water distribution in the slices. The slices pretreated with cyclic GMP at 0°C failed to decrease PAH accumulation. Cyclic GMP had no effect on organic cation (tetraethylammonium, TEA) transport in the slices. These evidence suggest that cyclic GMP may be able to modulate the rat renal transport system for PAH although this study does not explain the mechanism of inhibition of PAH transport by the cyclic nucleotide.

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A radioimmunoassay was developed for the measurement of exogenous thyrotropin-releasing hormone (TRH) in the whole blood of rats, beagle-dogs and humans. Even at low temperature, TRH is degraded so quickly in the whole blood that stabilization of TRH in the blood is necessary. For this purpose, the direct extraction with methanol and concentration before the radioimmunoassay was satisfactorily performed. The method is highly sensitive so that a lower quantifiable concentration of 20 pg/ml was detectable, and good reproducibility and standard errors of less than 10% from triplicate standard curves were obtained. At low concentration of TRH, the effect of food ingestion and volume of whole blood on the sensitivity of the radioimmunoassay was observed and thin layer chromatographic treatment improved it.

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1-(2-Tetrahydrofuryl)-5-fluorouracil (FT-207) concentrations in lymph and plasma were measured after rectal administration of three types of suppositories containing FT-207. The suppositories used in this study were prepared from polyacrylic acid aqueous gel bases, emulsion type bases and a commercial fat base, witepsol w-35. Both aqueous gel and emulsion type suppositories were superior to the suppositories fromwitepsol with regard to FT-207 concentration in lymph and plasma. The addition of aqueous gel to the polyacrylic acid aqueous gel was found to be effective for the lymph transport from the aqueous gels. Polyacrylic acid aqueous gel containing 1% glycyrrhizin was better than other suppositories. Solidification of the emulsion type bases are also investigated, and it is suggested that these may be better substitutes of the oily suppository.

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In the present study we investigated the relationship between the in vivo effects and the in vitro effects of serine and thiol proteinase inhibitors. Each one of the inhibitors, leupeptin, dansyl-L-leucyl-L-argininal (Dan-Leu-Argal), pyroglutamyl-L-leucyl-L-argininal (Pyr-Leu-Argal), E-64C or EP-459 was given intraperitoneally to mice for 8 consecutive days and various enzymatic activities were tested in 6 organs : forelimb muscle, hindlimb muscle, heart, spleen, liver and kidney. A multivariate analysis clearly differentiated the effects of leupeptin from those of its two analogues (Dan-Leu-Argal and Pyr-Leu-Argal), while the two analogues showed close relations with each other as to their in vivo effects. E-64C and EP-459 also showed resemblance of in vivo effects between them, but the effects of these two agents were clearly differentiated from those of leupeptin and its two analogues. This kind of study seems important to assess the in vivo effects of physiologically active substances including enzyme inhibitors.

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Anticonvulsant activities of diazepam in mice given pentylenetetrazol (PTZ), as well as effects of diazepam against potentiating activities of caffeine on these convulsions, were studied. Electroencephalographic (EEG) and electromyographic (EMG) recordings and behavioral observations were employed. First spike, clonic and tonic convulsions induced by PTZ were readily recognized from the EEF recording coupled with EMG recording. Pretreatment of mice with diazepam suppressed the incidence of clonic and tonic convulsions, but did not affect the development of first spike. These anticonvulsive effects of diazepam were also observed when PTZ-induced convulsions were potentiated with caffeine, except for clonic convulsion which was not suppressed with diazepam. The mechanisms of the antagonistic effects between diazepam and caffeine are discussed.

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The effects of tetrahydro-2-furanyl- and tetrahydro-2-pyranyl-ethers of (1→3)-β-D-glucans from Alcaligenes faecalis var.myxogenes (IFO 13140) on the activities of macrophages and natural killer (NK) cells in either ICR or BALB/c mice were studies. The derivatives with strong antitumor activity against Sarcoma 180 in ICR mice induced higher macrophage tumoricidal activity (>40%) than those with low antitumor activity (<20%). All derivatives with high or low antitumor activity augmented NK cell activity. On the other hand, the derivatives which show the high antitumor activity against Meth-A in BALB/c mice enhanced macrophage tumoricidal activity, while those with low or no antitumor activity did not. NK cell activity was activated by the derivatives with or without the antitumor activity. Furthermore, certain derivative which has high antitumor activity against Sarcoma 180 in ICR mice did not increase serum component (LB) in ICR mice. These results indicate that the antitumor activity of the derivatives correlates only with the capacity to cause macrophage actiation in vivo.

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The acidic fraction of bakers' yeast mannan containing mannose (93.6%), nitrogen (1.0%), and phosphorus (0.6%), designated as WAM025, showed a marked antitumor activity against the ascites forms of Ehrlich, sarcoma 180 and Meth A tumors in vivo. Namely, a 100% survival of mice transplanted with Ehrlich or sarcoma 180 ascites tumor, 1×10⁵ cells/ddY strain mouse, was obtained by the intraperitoneal administration of WAM025, 150 mg/kg/d for 10 d. This effect evoked by WAM025 seems to be attributed to the activation of peritoneal adherent cells in mice, because a significant elevation of lysosomal enzyme level and active oxygen generation was observed on the peritoneal adherent cells obtained from the mannan-administered mice.

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The present study provides the evidence that liver aldehyde oxidase in the presence of its electron donors can catalyze the reduction of N-hydroxyurethane ot urethane under anaerobic conditions. Guinea pig liver 9000×g supernatant and cytosol, but not liver microsomes, exhibited N-hydroxyurethane reductase activity in the presence of acetaldehyde or 2-hydroxypyrimidine. The cytosolic enzyme was precipitated with ammonium sulfate between 30 and 45% ammonium sulfate saturation. The N-hydroxyurethane reductase and aldehyde oxidase activities of the precipitate were similarly susceptible to inhibition by a variety of chemicals. When the precipitate was chromatographed on a DEAE-cellulose column, the elution peak position of N-hydroxyurethane reductase was entirely identical with that of aldehyde oxidase. Furthermore, purified rabbit liver aldehyde oxidase also exhibited a significant N-hydroxyurethane reductase activity in the presence of acetaldehyde or 2-hydroxypyrimidine.

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The dissolution profiles of furosemide in various solutions were studied with plain tablet and retard capsule of furosemide. The rate constant and percent dissolution in retard capsule were lower than that in plain tablet. Clinical pharmacokinetics and diuretic effect of furosemide after oral administration of two dosage forms were also studied with 3 normal subjects and 3 cirrhotic patients. In normal subjects, the extent of furosemide absorption from retard capsule was 45% of plain tablet. The daily urine volume after oral administration of two dosage forms was comparable, howerve, quite different porfiles were observed between these dosage forms. In patients, the extent of furosemide absorption in retard capsule was one half that of plain tablet. The dose of retard capsule was increased without adverse reactions, thus decreasing of ascites, which was not observed with plain tablet, was achieved with increasing urine volume.

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The preparation and antigenic properties of estriol 3-glucuronide-bovine serum albumin (BSA) conjugate in which the hapten is linked to the carrier protein through an (O-3-carboxypropylcarbamoylmethyl) oxime bridge at the C-6 position on the steroid nucleus, have been described. 16, 17-Di-O-acetyl-6-oxoestriol 6-(O-carboxymethyl) oxime 3-glucuronide acetate-methyl ester was condensed with γ-amino-n-butyric acid by the mixed anhydride method. Subsequent coupling with BSA followed by removal of the protecting groups with alkali gave the desired hapten-BSA conjugate. The antisera elicited in the rabbit with the conjugate were highly specific to estriol 3-glucuronide, discriminating from other 3-substituted compounds. The specificity of antisera improved by elongation of the 6-(O-carboxymethyl) oxime bridge has been discussed.

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