Mariann Zerbes

Mariann Zerbes; Stephen J. Bunn; David Powis

The characteristics and properties of the increase in cytosolic [Ca2+] that occurs in bovine adrenal medullary chromaffin cells on exposure to histamine have been investigated. Specifically, these experiments were conducted to determine how much external Ca2+ enters the cell through a (capacitative) Ca2+ entry pathway activated as a consequence of intracellular Ca2+ store mobilization, relative to that which enters independently of store depletion via other channels activated by histamine. In Fura-2 loaded cells continued exposure to histamine (10 microM) caused a rapid but transient increase in cytosolic [Ca2+] followed by a lower plateau that was sustained as long as external Ca2+ was present. In the absence of external Ca2+, only the initial brief transient was observed. In cells previously treated with thapsigargin (100 nM) in Ca(2+)-free medium to deplete the internal Ca2+ stores, histamine caused no increase in cytosolic [Ca2+] when external Ca2+ was absent. Re-introduction of external Ca2+ to thapsigargin-treated store-depleted cells caused a sustained increase in cytosolic [Ca2+] that was further increased (P < 0.0002) upon exposure to histamine. The histamine-evoked increase was prevented by the H1-receptor antagonist, mepyramine (2 microM). A comparison was made between store-dependent Ca2+ entry consequent upon store mobilization with histamine in Ca(2+)-free medium and plateau phase Ca2+ entry resulting from stimulation with histamine in Ca(2+)-containing medium. The latter was found to be approximately 3 times greater in magnitude than the former (P << 0.0001) at the same concentration of histamine (10 microM). It is concluded that histamine causes Ca2+ entry not only via a capacitative entry pathway secondary to internal store mobilization, but also causes substantial Ca2+ entry through other pathways.

D.B. De Laney; A.E. Jones; Mariann Zerbes; Gregory A. Tannock

Field samples were received for Mareks disease virus (MDV) isolation from clinically affected flocks from several regions of Eastern Australia. Lymphocytes were fractionated in Ficoll-Paque and passaged once or twice in chicken embryo kidney cultures. Serotype-specific virus was detected in infected cultures by indirect immunofluorescence with monoclonal antibodies. Serotype 1 MDV was isolated from 10 flocks. In samples from 5 of these flocks, serotype 2 and 3 vaccine viruses were isolated from the same specimen. In a parallel study, plasmas obtained during lymphocyte isolation were tested for antibodies to MDVs by agar gel precipitin (AGP) tests using serotype 1 and 3 antigen extracts. No correlation was observed between the rate of virus isolation and AGP positivity. The AGP test was incapable of discriminating between the different MDV serotypes.

Philip D. Marley; Peter J. R. Bales; Mariann Zerbes; David Powis; Melanie O'Farrell

Abstract: The effect on exocytosis of La3+, a known inhibitor of plasma membrane Ca2+‐ATPases and Na+/Ca2+ exchangers, was studied using cultured bovine adrenal chromaffin cells. At high concentrations (0.3‐3 mM), La3+ substantially increased histamine‐induced catecholamine secretion. This action was mimicked by other lanthanide ions (Nd3+, Eu3+, Gd3+, and Tb3+), but not several divalent cations. In the presence of La3+, the secretory response to histamine became independent of extracellular Ca2+. La3+ enhanced secretion evoked by other agents that mobilize intracellular Ca2+ stores (angiotensin II, bradykinin, caffeine, and thapsigargin), but not that due to passive depolarization with 20 mM K+. La3+ still enhanced histamine‐induced secretion in the presence of the nonselective inhibitors of Ca2+‐permeant channels SKF96365 and Cd2+, but the enhancement was abolished by prior depletion of intracellular Ca2+ stores with thapsigargin. La3+ inhibited 45Ca2+ efflux from preloaded chromaffin cells in the presence or absence of Na+. It also enhanced and prolonged the rise in cytosolic [Ca2+] measured with fura‐2 during mobilization of intracellular Ca2+ stores with histamine in Ca2+‐free buffer. The results suggest that the efficacy of intracellular Ca2+ stores in evoking exocytosis is enhanced dramatically by inhibiting Ca2+ efflux from the cell.

Peter J. R. Bales; Mariann Zerbes; David Powis; Philip D. Marley

The effects of Gd3+ on bradykinin‐ (BK‐) induced catecholamine secretion, 45Ca2+ efflux and cytosolic [Ca2+] were studied using bovine adrenal chromaffin cells. BK increased secretion in a Ca2+‐dependent manner. From 1–100 μM, Gd3+ progressively inhibited secretion induced by 30 nM BK to near‐basal levels, however from 0.3–3 mM Gd3+ dramatically enhanced BK‐induced secretion to above control levels. Gd3+ also increased basal catecholamine secretion by 2–3 fold at 1 mM. These effects were mimicked by Eu3+ and La3+. Gd3+ enhanced secretion induced by other agonists that mobilize intracellular Ca2+ stores, but simply blocked the response to K+. Gd3+ still enhanced basal and BK‐induced secretion in Ca2+‐free solution or in the presence of 30 μM SKF96365, however both effects of Gd3+ were abolished after depleting intracellular Ca2+ stores. Gd3+ (1 mM) reduced the rate of basal 45Ca2+ efflux by 57%. In Ca2+‐free buffer, BK transiently increased cytosolic [Ca2+] measured with Fura‐2. The [Ca2+] response to BK was substantially prolonged in the presence of Gd3+ (1 mM). The results suggest that Gd3+ greatly enhances the efficacy of Ca2+ released from intracellular stores in evoking catecholamine secretion, by inhibiting Ca2+ extrusion from the cytosol. This suggests that intracellular Ca2+ stores are fully competent to support secretion in chromaffin cells to levels comparable to those evoked by extracellular Ca2+ entry. Drugs that modify Ca2+ extrusion from the cell, such as lanthanide ions, will be useful in investigating the mechanisms by which intracellular Ca2+‐store mobilization couples to Ca2+‐dependent exocytosis.

Yvette Arvidson; Gregory A. Tannock; A. Senthilselvan; Mariann Zerbes

SummaryA model for determining immunogenic relationships between strains of infectious bronchitis virus is described that is based upon the vaccinating dose required to induce prevention of multiplication of a standard challenge dose of an homologous strain in the lungs of specific-pathogen-free chickens. The model has been used to demonstrate that:(1)Approximately 100 times the vaccinating dose must be used to produce immunity from one compared with two doses;(2)differences of immunogenicity of >1,000-fold exist between Australian strains;(3)immunogenicity is greatest for live virus administered directly to the trachea, followed by live virus administered by the ocular route, inactivated virus administered intratracheally and inactivated virus administered intramuscularly;(4)for chickens inoculated with Australian vaccine strains, protection against challenge is unrelated to serotype.

David Powis; Mariann Zerbes

Abstract: Reduction of intracellular Ca2+ stores (by chelation with the EDTA structural analogue, TPEN) is itself a sufficient trigger of capacitative Ca2+ entry. TPEN may be a useful tool in studies to investigate the mechanism of capacitative Ca2+ entry, since potential confounders, such as changes in cytosolic Ca2+ and second messenger levels, are obviated and the use of pharmacological modifiers of SERCA pumps and Ca2+ release channels is avoided.

David Powis; Mariann Zerbes; Lynn Herd; Peter R. Dunkley

The characteristics and properties of the increase in cytosolic [Ca2+] that occurs in bovine adrenal medullary chromaffin cells on exposure to angiotensin II have been investigated. In fura-2 loaded cells exposure to a maximally effective concentration of angiotensin II (100 nM) caused a rapid, but transient increase in cytosolic [Ca2+] followed by a lower plateau that was sustained as long as external Ca2+ was present. In the absence of external Ca2+ only the initial brief transient was observed. In cells previously treated with thapsigargin in Ca2+-free medium to deplete the internal Ca2+ stores, angiotensin II caused no increase in cytosolic [Ca2+] when external Ca2+ was absent. Reintroduction of external Ca2+ to thapsigargin-treated, store-depleted cells caused a sustained increase in cytosolic [Ca2+] that was not further increased upon exposure to angiotensin II. Analysis of the data suggests that in bovine chromaffin cells angiotensin II causes Ca2+ entry via a pathway(s) activated as a consequence of internal store mobilization, and entry through this pathway(s) forms the majority of the sustained Ca2+ influx evoked by angiotensin II.

Neil R. McGregor; Henry L. Butt; Mariann Zerbes; Iven Klineberg; R. H. Dunstan; Timothy K. Roberts

Henry L. Butt; R. H. Dunstan; Neil R. McGregor; Timothy K. Roberts; Mariann Zerbes; Iven Klineberg

Neil R. McGregor; R. Hugh Dunstan; Mariann Zerbes; Henry L. Butt; Timothy K. Roberts; Iven Klineberg