Molecular and functional characterization of gap junctions in the avian inner ear.

To analyze the fundamental role of gap junctions in the vertebrate inner ear, we examined molecular and functional characteristics of gap junctional communication (GJC) in the auditory and vestibular system of the chicken. By screening inner ear tissues for connexin isoforms using degenerate reverse transcription-PCR, we identified, in addition to chicken Cx43 (cCx43) and the inner-ear-specific cCx30, an as yet uncharacterized connexin predicted to be the ortholog of the mammalian Cx26. In situ hybridization indicated that cCx30 and cCx26 transcripts were both widely expressed in the cochlear duct and utricle in an overlapping pattern, suggesting coexpression of these isoforms similar to that in the mammalian inner ear. Immunohistochemistry demonstrated that cCx43 was present in gap junctions connecting supporting cells of the basilar papilla, in which its immunofluorescence colocalized with that of cCx30. However, cCx43 was absent from supporting cell gap junctions of the utricular macula. This variation in the molecular composition of gap junction plaques coincided with differences in the functional properties of GJC between the auditory and vestibular sensory epithelia. Fluorescence recovery after photobleaching, adapted to examine the diffusion of calcein in inner ear explants, revealed asymmetric communication pathways among supporting cells in the basilar papilla but not in the utricular macula. This study supports the hypothesis that the coexpression of Cx26/Cx30 is unique to gap junctions in the vertebrate inner ear. Furthermore, it demonstrates asymmetric GJC within the supporting cell population of the auditory sensory epithelium, which might mediate potassium cycling and/or intercellular signaling

Restoring the balance: regeneration of hair cells in the vestibular system of the inner ear

Loss of the sensory “hair cells” (HCs) from the vestibular (balance) system of the inner ear results in dizziness and balance dysfunction contributing to falls. In the inner ears of non-mammalian vertebrates, there is spontaneous and complete replacement of lost HCs. The regenerates derive from the non-sensory supporting cells (SCs) that surround each HC either from the daughter cells following SC division or by direct, non-mitotic conversion of SCs. In mammals, there is a very limited capacity to regenerate vestibular HCs but only a small percentage is replaced. They arise exclusively by SC conversion. Recent work in mice, and some in humans, has shown possibilities for inducing SCs to convert to cells expressing some HC characteristics, but that differentiation to fully functional HCs is incomplete. Identification of necessary transcription factors, and/or epigenetic modifiers as well as targets to promote SC proliferation is ongoing

Myosin VIIA is required for aminoglycoside accumulation in cochlear hair cells.

Myosin VIIA is expressed by sensory hair cells and has a primary structure predicting a role in membrane trafficking and turnover, processes that may underlie the susceptibility of hair cells to aminoglycoside antibiotics. [3H]Gentamicin accumulation and the effects of aminoglycosides were therefore examined in cochlear cultures of mice with different missense mutations in the myosin VIIA gene, Myo7a, to see whether myosin VIIA plays a role in aminoglycoside ototoxicity. Hair cells from homozygous mutant Myo7a(sh1) mice, with a mutation in a non-conserved region of the myosin VIIA head, respond rapidly to aminoglycoside treatment and accumulate high levels of gentamicin. Hair cells from homozygous mutant Myo7a(6J) mice, with a mutation at a highly conserved residue close to the ATP binding site of the myosin VIIA head, do not accumulate [3H]gentamicin and are protected from aminoglycoside ototoxicity. Hair cells from heterozygotes of both alleles accumulate [3H]gentamicin and respond to aminoglycosides. Although aminoglycoside uptake is thought to be via apical surface-associated endocytosis, coated pit numbers on the apical membrane of heterozygous and homozygous Myo7a(6J) hair cells are similar. Pulse-chase experiments with cationic ferritin confirm that the apical endocytotic pathway is functional in homozygous Myo7a(6J) hair cells. Transduction currents can be recorded from both heterozygous and homozygous Myo7a(6J) hair cells, suggesting it is unlikely that the drug enters via diffusion through the mechanotransducer channel. The results show that myosin VIIA is required for aminoglycoside accumulation in hair cells. Myosin VIIA may transport a putative aminoglycoside receptor to the hair cell surface, indirectly translocate it to sites of membrane retrieval, or retain it in the endocytotic pathway

Descartes, corpuscles and reductionism : mechanism and systems in Descartes' physiology

I argue that Descartes explains physiology in terms of whole systems, and not in terms of the size, shape and motion of tiny corpuscles (corpuscular mechanics). It is a standard, entrenched view that Descartes’s proper means of explanation in the natural world is through strict reduction to corpuscular mechanics. This view is bolstered by a handful of corpuscular-mechanical explanations in Descartes’s physics, which have been taken to be representative of his treatment of all natural phenomena. However, Descartes’s explanations of the ‘principal parts’ of physiology do not follow the corpuscular–mechanical pattern. Des Chene (2001) has identified systems in Descartes’s account of physiology, but takes them ultimately to reduce down to the corpuscle level. I argue that they do not. Rather, Descartes maintains entire systems, with components selected from multiple levels of organisation, in order to construct more complete explanations than corpuscular mechanics alone would allow

Characterizing human vestibular sensory epithelia for experimental studies: new hair bundles on old tissue and implications for therapeutic interventions in ageing.

Balance disequilibrium is a significant contributor to falls in the elderly. The most common cause of balance dysfunction is loss of sensory cells from the vestibular sensory epithelia of the inner ear. However, inaccessibility of inner ear tissue in humans severely restricts possibilities for experimental manipulation to develop therapies to ameliorate this loss. We provide a structural and functional analysis of human vestibular sensory epithelia harvested at trans-labyrinthine surgery. We demonstrate the viability of the tissue and labeling with specific markers of hair cell function and of ion homeostasis in the epithelium. Samples obtained from the oldest patients revealed a significant loss of hair cells across the tissue surface, but we found immature hair bundles present in epithelia harvested from patients >60 years of age. These results suggest that the environment of the human vestibular sensory epithelium could be responsive to stimulation of developmental pathways to enhance hair cell regeneration, as has been demonstrated successfully in the vestibular organs of adult mice

Nonattacking Queens in a Rectangular Strip

The function that counts the number of ways to place nonattacking identical chess or fairy chess pieces in a rectangular strip of fixed height and variable width, as a function of the width, is a piecewise polynomial which is eventually a polynomial and whose behavior can be described in some detail. We deduce this by converting the problem to one of counting lattice points outside an affinographic hyperplane arrangement, which Forge and Zaslavsky solved by means of weighted integral gain graphs. We extend their work by developing both generating functions and a detailed analysis of deletion and contraction for weighted integral gain graphs. For chess pieces we find the asymptotic probability that a random configuration is nonattacking, and we obtain exact counts of nonattacking configurations of small numbers of queens, bishops, knights, and nightriders.Comment: 21 pages, 3 figures, preprint of published version copyright Springer Basel AG 2011, Published online February 15, 2011, submitted March 15, 200

Multi-Year Application of Dairy Slurry on Grassland: Effects on Crop, Soil Biota, Soil Nutrients, and N\u3csub\u3e2\u3c/sub\u3eO Emission

The long-term effects of using manure as the principal nutrient source in intensive crop production systems are not well known. This paper reports on the effects of multi-year application of fertilizer or dairy slurry on a tall fescue (Festuca arundinacea Schreb.) sward. Slurry sustained greater grass yield than chemical fertilizer. Unlike fertilizer, slurry supplied 70 to 120 kg N/ha one year after application but little after one year; 4-years of manure applications built up the stable organic matter pool in the soil. Manure-N was less prone to leaching but more prone to N2O emissions than fertilizer-N. Manured soils had considerably more biological activity than fertilized soils. High rates of manure application increased soil P

Gentamicin alters membrane structure as shown by freeze-fracture of liposomes

Freeze-fracture has been used to examine the effects of gentamicin on membrane structure in liposomes of different anionic phospholipids combined with a neutral phospholipid, phosphatidylcholine. The molar ratios of neutral: anionic lipid were 1:1 (high anionic lipid ratio) and 4:1 (low anionic lipid) and the liposomes were incubated with 0.1 mM (low) and 1 mM (high) gentamicin. With the anionic phospholipid phosphatidylinositol bisphosphate, an identifiable disruption of the membrane bilayer was observed as well as aggregation of liposomes leading to membrane fusion. These effects occurred both at low gentamicin concentration and low anionic lipid content of the liposomes; these responses were not inhibited by 1 mM Ca2+. With the other anionic lipids tested (phosphatidylserine, phosphatidylinositol and phosphatidylinositol monophosphate), only aggregation and fusion of liposomes was observed and this effect only occurred at high gentamicin concentration and high anionic lipid content. Further, 1 mM Ca2+ inhibited the responses of these other anionic lipids to gentamicin. The results demonstrate the unique character of the interaction between gentamicin and phosphatidylinositol bisphosphate and provide further support for the hypothesis that a specific binding to this lipid is a key step in the ototoxic action of aminoglycoside antibiotics. They also suggest that such an interaction in vivo might cause alterations to the structure and properties of cell membranes in the inner ear.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/28139/1/0000591.pd