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Allatostatin (AST)-like immunoreactivity (IR) was localized in the eyestalk of Penaeus monodon by immunohistochemistry using four anti-AST antibodies. Depending on the antisera, AST-like immunoreactivity was detected in neuronal bodies of the lamina ganglionalis, cell bodies anterior to the medulla externa and cell bodies on the anterior and posterior of the medulla terminalis. Neuronal processes in neuropiles of the medulla externa, medulla terminalis, sinus gland and nerve fibers in the optic nerve were also recognized. No IR in cell bodies or in nerve fibers was found in the medulla interna. Strong AST-like immunoreactivity was found in hundreds of cells of the X organ. The localization of AST-like peptides suggests that they function as neurotransmitters and/or neuromodulators. Antiserum to the Drosophila AST receptor (Dar-2) recognized a single protein in P. monodon eyestalk protein extracts that was identical in size to that found in Drosophila protein extracts. Using this antiserum the putative P. monodon AST receptor was localized to the sinus gland in both juvenile and adult eyestalks. To our knowledge this is the first demonstration of a neuropeptide receptor localized to the crustacean sinus gland. This suggests that ASTs may function directly on the sinus gland as a neuromodulator. In juvenile eyestalks, the putative AST receptor was also localized to neuronal X organ cells of the medulla terminalis in males but not in females. The significance of this sex-specific receptor localization is unclear but emphasizes that ASTs function within the nervous system of the eyestalk.

Twenty-two isolates of Cymbidium mosaic virus (CyMV) were isolated from 35 orchid plants suspected of being infected with CyMV. Among the three methods used for detecting CyMV, immunoelectron microscopy (IEM-1) was shown to be the most sensitive method, being able to detect the virus in 71.43% of suspected CyMV-infected plants while the electron microscopic method and the indexing plant method could detect 51.43 and 42.86%, respectively. Out of 12 symptomless plants investigated, 25% were found by IEM-1 method to be infected with the virus. Purified CyMV were flexuous rods having lengths between 470-490 nm. A few end-to-end aggregates were also observed and the 280 260 absorbance ratios were from 0.884 to 0.929. The yield of CyMV was 31.07 to 44.09 mg per kg of Datura leaves. Antibodies against purified CyMV D2 were produced in rabbits and hens. The antibody titers in the yolk and sera of hens indicated that 0.5 mg of virus per immunization efficiently generated an abundant supply of IgY in the yolk, however 1 mg of virus per immunization gave a stronger immune response in both sera and yolk. The average yields of IgY were 6.5 +/- 0.6 and 9.4 +/- 0.9 mg/ml of yolk in the group that received 0.5 mg and the group that received 1.0 mg of the virus, respectively. Positive ELISA reactions were observed in 18 and 20 of 22 CyMV isolates when detected with rabbit IgG and IgY, respectively, demonstrating that those isolates were serologically related and the ELISA reactions were shown to be stronger with IgY than those with rabbit IgG in most isolates. The degree of reaction between the CyMV isolates, O(2) and O(4), and IgY was less than that of the other isolates. The two isolates, D(6) and Cat(6), gave negative reactions to rabbit IgG. The results of ELISA assays showed that the homologous serological reaction was not consistently stronger than the heterologous one. Twelve isolates out of twenty-two gave stronger reactions than the homologous antigen (CyMV D(2)) when IgY was used as the detecting antibody while nine isolates gave stronger reactions when using rabbit IgG. No reactions were observed with other plant viruses and plant proteins from healthy Datura.

This study defined structural requirements for organophosphorous and methylcarbamate insecticides for altering the L-kynurenine pathway of L-tryptophan metabolism in mice. Kynurenine formamidase inhibition by organophosphorous acid triesters and methylcarbamates is the proposed primary event resulting in increase in xanthurenic acid urinary excretion and plasma L-kynurenine. Alteration of the L-kynurenine pathway occurred with compounds that inhibited liver kynurenine formamidase by more than 80%. Pyrimidinyl phosphorothioates followed by crotonamide phosphates were the most potent compounds that changed L-tryptophan metabolism, i.e., pirimiphos-ethyl (20 mg/kg) inhibited liver kynurenine formamidase by 99%, and increased xanthurenic acid urinary excretion and plasma L-kynurenine by 576 +/- 195 and 330 +/- 44%, respectively. Replacement of sulphur by oxygen in the phosphorothioate diazinon reduced in vivo liver kynurenine formamidase inhibition. Consequently, xanthurenic acid urinary excretion and plasma L-kynurenine were not elevated. Atropine, cycloheximide, 2-PAM and phenylmethylsulfonyl fluoride did not alleviate diazinon-altered L-tryptophan metabolism. Because of the potential of the majority of organophosphorous acid triesters and methylcarbamates to inhibit kynurenine formamidase, this novel noncholinergic mechanism warrants consideration in assessment of organophosphorous and methylcarbamate toxicity in occupational and accidental exposures.

It was previously demonstrated that a correlation existed between estrogen receptor (ER) status and levels of estrogen sulfotransferase in human mammary cancer, high levels being associated with ER-positive and low levels with ER-negative tumors. We have now examined these same parameters, along with progesterone receptor (PGR) status, in 44 primary human mammary tumors. Tumors which were both ER-positive and PGR-positive (n = 21), showed significantly higher levels of estrogen sulfotransferase compared to ER-positive PGR-negative (n = 12) and ER-negative PGR-negative (n = 11) tumors. These values (pmol estradiol sulfate per mg protein per 2 hr) were 72 +/- 12 (S.E.), 25 +/- 5, and 11 +/- 4 (p less than 0.01 and less than 0.005, respectively). There was no significant difference between ER-positive PGR-negative and ER-negative PGR-negative tumors. The possible involvement of PGR in the regulation of estrogen sulfotransferase is discussed.

Estrogen receptor (ER) levels, formation of 3'-phosphoadenosine-5'phosphosulfate (PAPS), and sulfate transfer to estradiol-178, have been determined in 62 human primary carcinoma cytosol preparations. Whilst no differences between ER positive and ER negative tumors were found in the synthesis of [35S]PAPS, formation of estradiol-3-[35S]sulfate catalysed by estrogen sulfotransferase was significantly higher in ER positive tumors [84 +/- 10 (mean +/- S.E.M.) versus 32 +/- 9 pmole estradiol-3-sulfate per mg protein per 2 hr for ER positive and ER negative tumors, respectively, p less than 0.02]. By choosing a discriminating value for estrogen sulfurylation of 40 pmole per mg protein per 2 hr, then 16/19 (84%) of the ER negative tumors were below this point. Since previous data had shown that 20/23 (87%) progesterone receptor negative tumors were also below this value (Pewnim, Adams and Ho, CANCER RES. 40, 1360 (1980), an alternative simple method, having a high potential for the evaluation of hormone responsiveness in mammary cancer, is suggested.

A positive correlation between the expression of estrogen sulphotransferase (EC 2.8: 2.4) and the estrogen receptor (ER) in human breast cancer tissues was previously demonstrated. We have now established that a similar correlation exists between the expression of hydroxysteroid sulphotransferase (EC 2.8: 2.2) and ER in such tissues. Enzyme activity was present in 93% of the ER + tumor cytosols (mean 59 +/- 44 (SD) pmol dehydroepiandrosterone sulphate formed per mg protein per 2 h (n = 42). Activity was detected in 68% of ER - tumors and this was significantly lower (mean 21 +/- 26 (SD) (n = 19), P less than 0.001) than the former group. Metabolism of estradiol-17 beta (E2) and the adrenal-derived estrogen 5-androstene-3 beta, 17 beta-diol (ADIOL), which is a substrate for hydroxysteroid sulphotransferase but not estrogen sulphotransferase, was studied in four ER + human mammary cancer cell lines (MCF-7, T47-D, MDA-MB-361 and ZR-75-1) and four ER-human mammary cell lines (BT-20, MDA-MB-231, MDA-MB-330 and HBL-100), employing steroid concentrations of 1 nM. At this concentration, formation of ester sulphates was a major route of metabolism in the ER + cell lines; E2 yielding a mean of 6.5 pmol estrogen monosulphates/mg DNA in 16 h and ADIOL yielding a mean of 9.4 pmol C19-5-ene steroid monosulphates/mg DNA in 16 h. In three of the four ER - cell lines, formation of sulphates from E2 occurred at an eight-fold lower rate (mean 0.8 pmol estrogen sulphates/mg DNA in 16 h), whereas MDA-MB-330 cells did not form estrogen sulphates. Only one of the four ER- cell lines (BT-20) sulphurylated ADIOL and this was at a 12-fold lower rate compared to the mean value for the ER + cel lines. Oxidation of E2 and ADIOL occurred in all cell lines and was generally the major route of metabolism in the ER - cells. A significant correlation between formation of estrone and dehydroepiandrosterone occurred for all cell lines (r = 0.98, P less than 0.001) indicating that the same 17 beta-hydroxysteroid dehydrogenase was probably involved. Since ADIOL is estrogenic in a number of systems at the concentration found in the blood of Western women (approximately 2 nM), the coordinated expression of hydroxysteroid sulphotransferase, estrogen sulphotransferase, and ER, supports the concept of a functional relationship between estrogen action via ER and sulphurylation reactions.

Estrogen sulfotransferase (EC 2.8.2.4) activity and estrogen receptor levels were measured in 32 human primary breast cancer cytosol preparations. Two types of tumors were identified: type 1, in which estrogen sulfotransferase levels were low (less than 40 pmol 17 beta-estradiol 3-sulfate formed per mg protein per 2 hr) and were independent of [35S]adenosine 3'-phosphate 5'-phosphosulfate production from [35S]sulfate and adenosine triphosphate, and type 2, in which estrogen sulfotransferase levels ranged from 50 to 200 pmol 17 beta-estradiol 3-sulfate per mg protein per 2 hr and were correlated with [35S]adenosine 3'-phosphate 5'-phosphosulfate formation (r = 0.70; p less than 0.005). In type 1 tumors, 11 of 16 were estrogen receptor negative; in type 2 tumors, 2 of 16 were receptor negative. Estrogen sulfotransferase levels in receptor-negative tumors were significantly lower than the levels in receptor-positive tumors (p = 0.025).