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Different aspects of the mode of action of cytokinin in the moss Physcomitrella patens were studied. In the search for a cytokinin receptor, a particulate fraction was isolated which had cytokinin-binding activity . Cytokinin-binding a c t ivi ty is described as the amount of binding of a radioactive cytokinin that is prevented from binding by excess unlabel led cytokinin. The cytokinin-binding activity was only detected with a tritiated cytokinin of high specific activity. Two assays were used, an equil ib r ium d ia ly s i s and a c ent r i fugat ion assay. These assays were used to show that the cytokinin-binding a c t iv i ty was heatlab i le and can be solubilised by the detergent Triton X100, but not by acetone. Cytokinin-binding a c t iv i ty was higher in phosphate-starved t is sue which consists mainly of caulonemata, the target cells for cytokinin action. A 13 ,0 0 0-8 0 ,000g pel let appeared to contain a component which has some of the characteristic s expected for a cytokinin receptor that was a membrane bound protein. Mutagenised spores were screened for cytokinin nonresponding mutants. No such mutants were obtained from 25,000 plants. A temperature-sensitive mutant, ove 409, was isolated which produced normal leafy shoots at I7°C, but many abnormal buds at 2 4 ° C . T h i s mutant was found to over-produce isopentenyladenine at the higher temperature. The level of cytokinin production in wild type and ove 78 was also found to be temperature-dependent. The change in cytokinin production by ove 409 was over twice that with wild type and ov.e 78, and dropped to wi ld type levels at 17°C. By the use of protoplast fusion, ove 409 was found to be recessive to wild type and in the same complementation group as ove A78. The role of light was studied in the induction of the gametophore. In the presence of supplied cytokinin, bud induction was caused by red light around 613 to 687nm. When the cytokinin-induced bud induction occurs in the dark after exposure to red light , bud induction is reduced by a short exposure to far-red light or a dark period before the addit ion of the cytokinin- This indicates that phytochrome is involved in bud induction. However, another factor appears to be involved in bud induction as this was greater when the cytokinin was applied to the cultures 2 hours after the exposure to red light. The possible role of calcium ions in bud induction was investigated. Calcium ions have been reported to induce the first stages of buds in the moss Funaria hygrometrica. Using the calcium ionophore A23187, calcium ions were shown to induce chloronemal branching in Physcomitre 11a patens. Therefore calcium ions appear to have a role in branching, but not in the differentiation of a bud.

Graduate

Ph.D. / The sensitivity of Petunia hybrida flowers to the plant hormone ethylene increases during the early stages of senescence and directly after pollination of the flowers. During these early stages .of senescence, ethylene production is very low and only increases after ethylene sensitivity had already increased. The increase in ethylene sensitivity is due to the forming of short-chain saturated fatty acids that accumulate in the petal tissue during the early stage of senescence. Following pollination, these acids are formed in the stylar tissue and transported to the corolla where it causes a rapid increase in ethylene sensitivity. The synthesis of short-chain saturated fatty acids is controlled by the enzyme acetyl-GoA carboxylase. As the acitivity of this enzyme decreases, the concentrations of short-chain fatty acids increase due to the lack of malonyl-GoA required for chain lengthening. During the early stages of senescence, the activity of this enzyme decreases simultaneously with an increase in short-chain fatty acids and ethylene-sensitivity. The results of this study indicate that acetyl-GoA carboxylase is a key enzyme involved in the control of ethylene sensitivity in petunia flowers.

It has been recorded that women respond to tricyclic antidepressives with a greater incidence of anticholinergic side effects than men do, particularly women taking an exogenous source of oestrogen. The aim of this study was to investigate the influence that ethinyl oestradiol and Premarin© had on the metabolism of a number of tricyclic antidepressives, and also the influence they had on the binding ability of microsomes to imipramine. Rat hepatocyctes and microsomes were used. Detection techniques used were High Pressure Liquid Chromatography and Spectrophotometry respectively. In addition to these studies, a study of the anticholinergic activity of Nomifensine, tricyclic antidepressives and their derivatives was performed on a rat jujenum. Results conclusively showed that ethinyl oestradiol had a marked influence on the metabolism of the tricyclic antidepressives studied. Premarin© had Iittle, if any influence. However, both ethinyl oestradiol and Premarin© affected the binding of microsomes to imipramine, but ethinyl oestradiol had the greater effect. The parent compound in each case exhibited a higher pAZ value. Results indicate that a possible explanation for the increased anticholinergic side effect is due to an inhibition of the metabolism of the tricyclic antidepressives by oestrogen.

18-hydroxycorticosterone, aldosterone and corticosterone were positively characterized in duck adrenal incubates. A compound resembling 11-dehydrocorticosterone was also present. The addition of 1 I.U. of ACTH/100 mg of adrenal tissue caused an increase in the production rate of all four steroids. The most dramatic effect however, was observed with respect to the synthesis of corticosterone. Aldosterone-4-¹⁴C and 18-hydroxycorticosterone-4-¹⁴C were isolated from the media of adrenal glands incubated in the presence of a tracer amount of corticosterone-4-¹⁴C. During the incubation the maximum specific activity recorded for aldosterone-4-¹⁴C was 66% of the specific activity of the precursor. This observation indicated that the primary pathway to aldosterone in the duck adrenal was probably via corticosterone. 18-hydroxycorticosterone may be an intermediate between corticosterone and aldosterone or may be a distinct end product of unknown biological activity. No difference in the biological half-life (T₁/2) of corticosterone was found between fresh-water maintained fresh-water loaded birds and birds maintained on fresh water or saline and loaded with saline. An increase in the apparent volume of distribution (A.V.D.) of corticosterone was found however in saline loaded ducks. This resulted in an increased secretory rate of corticosterone in those animals loaded with saline. In the saline maintained ducks an increased adrenal size was responsible for the enhanced corticosterone secretory rate. In those ducks maintained on fresh water and loaded with saline a weight specific increase in the corticosterone secretory rate was apparent. The implications of the increased corticosterone secretory rate are discussed in relation to the known dependence of nasal gland function in marine birds on a functional adrenal cortex. / Science, Faculty of / Zoology, Department of / Graduate

The effects of corticosterone, cortisol, aldosterone, oxytocin and vasopressin on the renal excretory pattern of the water-loaded domestic duck (Anas platyrhynchos) were examined. When compared with the results obtained from mammalian studies, the excretory pattern differed little with respect to water and sodium. The domestic duck showed high levels of potassium excretion. It is suggested that the function of the potassium secretory mechanism in the distal tubule may be predominant in this species. / Science, Faculty of / Zoology, Department of / Graduate

The purpose of this investigation was to determine whether or not the adrenocortical steroids influence the activity of the avian supra-orbital gland. In the intact saline loaded duck (Anas platyrhynchus), a diphasic excretory pattern was apparent, as an urinary diuresis was followed by activation of the supra-orbital glands. Variations in the circulating levels of adrenocortical steroids resulted in qualitative and quantitative changes in both phases of this response. Injection of exogenous, cortisol, cortexone and aldosterone caused an initial increase in the rate of nasal discharge and a consequent increase in the total output of Na⁺ and K⁺. Conditions of selective adrenocortical activity, i.e., subtotal adrenalectomy, severely impaired the extra-renal response while total adrenalectomy completely obliterated the response to an osmotic stimulus. The activity of the supra-orbital gland in the totally adrenalectomized duck was restored after treatment with cortisol. The renal phase in ducks treated with adrenocortical steroids was modified when compared to the control response. Administration of cortisol prior to saline loading resulted in a significantly lower total output of sodium although the volume of urine excreted did not differ from the control value. Treatment with aldosterone resulted in a significantly lower renal output of both Na⁺ and K⁺ as well as a decrease in urine volume. Cortexone and ACTH showed the control renal pattern of excretion. Unilateral adrenalectomy caused no apparent change in the renal excretory pattern whilst total adrenalectomy caused a significant increase in the total volume of urine excreted as well as on increase in the total output of Na⁺ and K⁺. Totally adrenalectomized birds maintained on cortisol reverted to a normal excretory pattern. The influence of adrenocortical steroids on the control of the avian supra-orbital gland is discussed. It can be said with certainty that adrenocortical steroids influence the activity of the supra-orbital gland of the Domestic Duck (Anas platyrhynchus). Furthermore, cortisol administered to the adrenalectomized animal reactivates the gland suggesting that a glucocorticoid is essential for its function. / Science, Faculty of / Zoology, Department of / Graduate