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Biomedical subjects

Stephen T Brown

Publications and source records attributed to Stephen T Brown.

9 recordsLinked to original sources
PubMed2005

Distinct roles for hindbrain and paraxial mesoderm in the induction and patterning of the inner ear revealed by a study of vitamin-A-deficient quail.

The hindbrain and cranial paraxial mesoderm have been implicated in the induction and patterning of the inner ear, but the precise role of the two tissues in these processes is still not clear. We have addressed these questions using the vitamin-A-deficient (VAD) quail model, in which VAD embryos lack the posterior half of the hindbrain that normally lies next to the inner ear. Using a battery of molecular markers, we show that the anlagen of the inner ear, the otic placode, is induced in VAD embryos in the absence of the posterior hindbrain. By performing grafting and ablation experiments in chick embryos, we also show that cranial paraxial mesoderm which normally lies beneath the presumptive otic placode is necessary for otic placode induction and that paraxial mesoderm from other locations cannot induce the otic placode. Two members of the fibroblast growth factor family, FGF3 and FGF19, continue to be expressed in this mesodermal population in VAD embryos, and these may be responsible for otic placode induction in the absence of the posterior hindbrain. Although the posterior hindbrain is not required for otic placode induction in VAD embryos, the subsequent patterning of the inner ear is severely disrupted. Several regional markers of the inner ear, such as Pax2, EphA4, SOHo1 and Wnt3a, are incorrectly expressed in VAD otocysts, and the sensory patches and vestibulo-acoustic ganglia are either greatly reduced or absent. Exogenous application of retinoic acid prior to 30 h of development is able rescue the VAD phenotype. By performing such rescue experiments before and after 30 h of development, we show that the inner ear defects of VAD embryos correlate with the absence of the posterior hindbrain. These results show that induction and patterning of the inner ear are governed by separate developmental processes that can be experimentally uncoupled from each other.

Animals↗
PubMed2005

Hypoxic augmentation of Ca2+ channel currents requires a functional electron transport chain.

The incidence of Alzheimer disease is increased following ischemic episodes, and we previously demonstrated that following chronic hypoxia (CH), amyloid beta (Abeta) peptide-mediated increases in voltage-gated L-type Ca(2+) channel activity contribute to the Ca(2+) dyshomeostasis seen in Alzheimer disease. Because in certain cell types mitochondria are responsible for detecting altered O(2) levels we examined the role of mitochondrial oxidant production in the regulation of recombinant Ca(2+) channel alpha(1C) subunits during CH and exposure to Abeta-(1-40). In wild-type (rho(+)) HEK 293 cells expressing recombinant L-type alpha(1C) subunits, Ca(2+) currents were enhanced by prolonged (24 h) exposure to either CH (6% O(2)) or Abeta-(1-40) (50 nm). By contrast the response to CH was absent in rho(0) cells in which the mitochondrial electron transport chain (ETC) was depleted following long term treatment with ethidium bromide or in rho(+) cells cultured in the presence of 1 microm rotenone. CH was mimicked in rho(0) cells by the exogenous production of O2(-.). by xanthine/xanthine oxidase. Furthermore Abeta-(1-40) enhanced currents in rho(0) cells to a degree similar to that seen in cells with an intact ETC. The antioxidants ascorbate (200 microm) and Trolox (500 microm) ablated the effect of CH in rho(+) cells but were without effect on Abeta-(1-40)-mediated augmentation of Ca(2+) current in rho(0) cells. Thus oxidant production in the mitochondrial ETC is a critical factor, acting upstream of amyloid beta peptide production in the up-regulation of Ca(2+) channels in response to CH.

Alzheimer Disease↗
PubMed2005

Dlx gene expression during chick inner ear development.

Members of the Dlx gene family play essential roles in the development of the zebrafish and mouse inner ear, but little is known regarding Dlx genes and avian inner ear development. We have examined the inner ear expression patterns of Dlx1, Dlx2, Dlx3, Dlx5, and Dlx6 during the first 7 days of chicken embryonic development. Dlx1 and Dlx2 expression was seen only in nonneuronal cells of the cochleovestibular ganglion and nerves from stage 21 to stage 32. Dlx3 marks the otic placode beginning at stage 9 and becomes limited to epithelium adjacent to the hindbrain as invagination of the placode begins. Dlx3 expression then resolves to the dorsal otocyst and gradually becomes limited to the endolymphatic sac by stage 30. Dlx5 and Dlx6 expression in the developing inner ear is first seen at stages 12 and 13, respectively, in the rim of the otic pit, before spreading throughout the dorsal otocyst. As morphogenesis proceeds, Dlx5 and Dlx6 expression is seen throughout the forming semicircular canals and endolymphatic structures. During later stages, both genes are seen to mark the distal surface of the forming canals and display expression complementary to that of BMP4 in the vestibular sensory regions. Dlx5 expression is also seen in the lagena macula and the cochlear and vestibular nerves by stage 30. These findings suggest important roles for Dlx genes in the vestibular and neural development of the avian inner ear.

Amino Acid Sequence↗
PubMed2004

Amyloid beta peptides mediate physiological remodelling of the acute O2 sensitivity of adrenomedullary chromaffin cells following chronic hypoxia.

OBJECTIVE: The non-neurogenic response of the neonatal adrenal medulla is vital in cardiovascular and respiratory development and to the survival of newborns exposed to hypoxic stress. Here, we examined the acute hypoxic response of immortalised rat adrenomedullary chromaffin cells following exposure to chronic hypoxia (CH; 6% O(2) for 24 h). METHODS: Ca(2+) and K(+) channel currents were recorded using by whole-cell patch-clamp. RESULTS: Following incubation in CH, the acute O(2) sensitivity of K(+) current in immortalised adrenomedullary chromaffin (MAH) cells was enhanced due to a selective increase in the density of an O(2)-sensitive Ca(2+)-dependent K(+) current, secondary to ROS-mediated augmentation of voltage-gated Ca(2+) currents. The effect of CH on Ca(2+) currents was not additive to exogenous Abeta(1-40) and was blocked by the gamma-secretase inhibitors gamma-X and gamma-VI, demonstrating a role for amyloid beta peptide (AbetaP) production. Ca(2+) current enhancement was abolished in the presence of the transcription inhibitor actinomycin D but unaffected by the vacuolar H(+) ATPase inhibitor bafilomycin A1. CONCLUSION: AbetaP production and transcriptional regulation during CH regulated the properties of a peripheral chemosensory cell, defining a role for these enigmatic peptides in the signalling pathway of a physiological response to CH in the developing cardiovascular system.

Amyloid beta-Peptides↗
PubMed2004

Acute and chronic hypoxic regulation of recombinant hNa(v)1.5 alpha subunits.

Acute and chronic hypoxic regulation of ion channels is involved in both cell physiology and pathology. Voltage-dependent Na(+) channels play a dominant role in the upstroke of the action potential in excitable cells, while non-inactivating (persistent or sustained) Na(+) currents contribute to action potential shape and duration. In cardiac myocytes, hypoxic augmentation of persistent Na(+) currents has been proposed to underlie cardiac arrhythmias via prolonging action potential duration. Here, we demonstrate that acute hypoxia (P(O2), 20mm Hg) augmented persistent Na(+) currents in HEK293 cells stably expressing human Na(v)1.5 alpha subunits. Hypoxia also inhibited peak Na(+) currents in a voltage-dependent manner, and the kinetics of activation and inactivation of Na(+) currents were significantly slowed during hypoxia. We further demonstrate that exposure to chronic hypoxia (6% O(2) for 24h) augmented peak Na(+) channel current, which given the exogenous promoter driving expression of the channel occurs most probably via a post-transcriptional mechanism. These effects of acute and chronic hypoxia likely play an arrhythmogenic role during both short- and long-term hypoxic/ischaemic episodes. The HEK293 expression system provides a useful paradigm in which to examine the mechanisms of O(2) sensing by the Na(+) channel.

Cell Hypoxia↗
PubMed2004

H2O2 regulates recombinant Ca2+ channel alpha1C subunits but does not mediate their sensitivity to acute hypoxia.

Acute hypoxic inhibition of the pore-forming alpha(1C) subunit of the L-type Ca(2+) channel mediates hypoxic arterial vasodilatation, a physiological response which matches tissue O(2) demand and supply in the systemic vasculature. In numerous O(2)-sensing cell types, reactive O(2) species (ROS) have been proposed as mediators linking lowered O(2) levels with the appropriate cellular response. In this study, we examined the roles of H(2)O(2) and NADPH oxidase as mediators of hypoxic inhibition of recombinant alpha(1C) subunits. Human cardiac L-type Ca(2+) channel alpha(1C) subunits were stably expressed in HEK 293 cells. Ca(2+) currents were recorded using the whole-cell configuration of the patch-clamp technique. Bath application of 100microM H(2)O(2) significantly enhanced depolarisation-evoked Ca(2+) currents in a voltage-dependent manner, while dialysis with 1000Uml(-1) catalase reduced these currents. In the presence of catalase, hypoxic inhibition of Ca(2+) currents was not significantly different compared to non-dialysed controls. The NADPH oxidase inhibitors diphenylene iodonium (10microM) and phenylarsine oxide (5microM) were without effect on either basal Ca(2+) currents or responses to hypoxia. Thus, endogenous production of H(2)O(2) regulates the alpha(1C) subunit. However, neither suppression of H(2)O(2) levels nor inhibition of NADPH oxidase is involved in O(2)-dependent regulation of the Ca(2+) channel.

Calcium Channels, L-Type↗