Pattern formation in the basilar papilla: evidence for cell rearrangement.

The avian basilar papilla is composed of hair and supporting cells arranged in a regular pattern in which the hair cells are surrounded and isolated from each other by supporting cell processes. This arrangement of cells, in which the apical borders of hair cells do not contact one another, may be generated by contact-mediated lateral inhibition. Little is known, however, about the way in which hair and supporting cells are organized during development. Whole mounts double-labeled with antibodies to the 275 kDa hair-cell antigen and the tight junction protein cingulin were therefore used to examine the development of cell patterns in the basilar papilla. Hair cells that contact each other at their apical borders are seen during early development, especially on embryonic days (E) 8 and 9, but are no longer observed after E12. Hair and supporting cell patterns were analyzed in three different areas of the papilla at E9 and E12. In two of these regions between E9 and E12, the ratio of supporting cells to hair cells does not change significantly, whereas there is an increase in both the number of supporting cells around each hair cell and the number of hair cells that each supporting cell contacts. In the third region examined, there is a dramatic rise in the number of supporting cells around each hair cell, which although accompanied by a small, significant increase in the ratio of supporting cells to hair cells cannot be accounted for by an increase in supporting cell numbers. These data show that a rearrangement of hair and supporting cells with respect to one another may be a fundamental process underlying the development of a regular pattern in the basilar papilla

A Novel Antigen Sensitive to Calcium Chelation That is Associated with the Tip Links and Kinocilial Links of Sensory Hair Bundles

Tip links are extracellular, cell-surface-associated filaments of unknown molecular composition that are thought to gate the mechanotransducer channel of the sensory hair cell. They disappear from the hair bundle in response to calcium chelation and lanthanum treatment and resist degradation by the protease subtilisin. A monoclonal antibody derived from a hybridoma screen identified a novel antigen associated with tip links, the tip-link antigen. The tip-link antigen is also associated with kinocilial links, subtilisin-resistant filaments that are sensitive to calcium chelation and connect the kinocilium to the tallest stereocilia of the hair bundle. Furthermore, the tip-link antigen is expressed in the retina, where it is associated with the ciliary calyx, a ring of microvilli that surrounds the outer segment of the photoreceptor. The tip-link antigen rapidly disappears from the surface of the hair bundle in response to calcium chelation. It is also subtilisin resistant, relative to the ankle-link antigen, an antigen associated with another type of hair bundle link. The tip-link antigen is lanthanum sensitive and, like tip links, reappears on the surface of the hair bundle after calcium chelation. The monoclonal antibody to the tip-link antigen immunoprecipitates two concanavalin A-reactive polypeptides with apparent molecular masses of 200 and 250 kDa from detergent extracts of the retina. These results provide the first identification of a cell surface antigen associated with tip links, indicate that tip links share properties in common with kinocilial links, and reveal a second epitope that, along with the ankle-link antigen, is common to both sensory hair bundles and the ciliary calyx of photoreceptors

A comparison of cortical and trabecular bone from C57 Black 6 mice using Raman spectroscopy

Peer reviewedPostprin

Effects of deleting cannabinoid receptor-2 on mechanical and material properties of cortical and trabecular bone

Acknowledgements We thank Dr J.S. Gregory for assistance with Image J and Mr K. Mackenzie for assistance with Micro-CT analysis. Funding ABK was funded by a University of Aberdeen, Institute of Medical Sciences studentship and the Overseas Research Students Awards Scheme.Peer reviewedPublisher PD

Mechanical and material properties of cortical and trabecular bone from cannabinoid receptor-1-null (Cnr1-/-) mice

Funding ABK was funded by a studentship from the University of Aberdeen, Institute of Medical Sciences, and the Overseas Research Students Awards Scheme Acknowledgments We are grateful to Dr J.S. Gregory for assistance with Image J and Mr K. Mackenzie for assistance with Micro-CT analysis.Peer reviewedPostprin

Stereociliary Myosin-1c Receptors Are Sensitive to Calcium Chelation and Absent from Cadherin 23 Mutant Mice

The identities of some of the constituents of the hair-cell transduction apparatus have been elucidated only recently. The molecular motor myosin-1c (Myo1c) functions in adaptation of the hair-cell response to sustained mechanical stimuli and is therefore an integral part of the transduction complex. Recent data indicate that Myo1c interacts in vitro with two other molecules proposed to be important for transduction: cadherin 23 (Cdh23), a candidate for the stereociliary tip link, and phosphatidylinositol 4,5-bisphosphate (PIP2), which is abundant in the membranes of hair-cell stereocilia. It is not known, however, whether these interactions occur in hair cells. Using an in situ binding assay on saccular hair cells, we demonstrated previously that Myo1c interacts with molecules at stereociliary tips, the site of transduction, through sequences contained within its calmodulin (CaM)-binding neck domain, which can bind up to four CaM molecules. In the current study, we identify the second CaM-binding IQ domain as a region of Myo1c that mediates CaM-sensitive binding to stereociliary tips and to PIP2 immobilized on a solid support. Binding of Myo1c to stereociliary tips of cochlear and vestibular hair cells is disrupted by treatments that break tip links. In addition, Myo1c does not bind to stereocilia from mice whose hair cells lack Cdh23 protein despite the presence of PIP2 in the stereociliary membranes. Collectively, our data suggest that Myo1c and Cdh23 interact at the tips of hair-cell stereocilia and that this interaction is modulated by CaM

Aminoglycoside-Induced Phosphatidylserine Externalization in Sensory Hair Cells Is Regionally Restricted, Rapid, and Reversible

The aminophospholipid phosphatidylserine (PS) is normally restricted to the inner leaflet of the plasma membrane. During certain cellular processes, including apoptosis, PS translocates to the outer leaflet and can be labeled with externally applied annexin V, a calcium-dependent PS-binding protein. In mouse cochlear cultures, annexin V labeling reveals that the aminoglycoside antibiotic neomycin induces rapid PS externalization, specifically on the apical surface of hair cells. PS externalization is observed within ~75 s of neomycin perfusion, first on the hair bundle and then on membrane blebs forming around the apical surface. Whole-cell capacitance also increases significantly within minutes of neomycin application, indicating that blebbing is accompanied by membrane addition to the hair cell surface. PS externalization and membrane blebbing can, nonetheless, occur independently. Pretreating hair cells with calcium chelators, a procedure that blocks mechanotransduction, or overexpressing a phosphatidylinositol 4,5-biphosphate (PIP2)-binding pleckstrin homology domain, can reduce neomycin-induced PS externalization, suggesting that neomycin enters hair cells via transduction channels, clusters PIP2, and thereby activates lipid scrambling. The effects of short-term neomycin treatment are reversible. After neomycin washout, PS is no longer detected on the apical surface, apical membrane blebs disappear, and surface-bound annexin V is internalized, distributing throughout the supranuclear cytoplasm of the hair cell. Hair cells can therefore repair, and recover from, neomycin-induced surface damage. Hair cells lacking myosin VI, a minus-end directed actin-based motor implicated in endocytosis, can also recover from brief neomycin treatment. Internalized annexin V, however, remains below the apical surface, thereby pinpointing a critical role for myosin VI in the transport of endocytosed material away from the periphery of the hair cell

A Clean, Green New Zealand? An In-Depth Look at the Personal Experiences of Animal Rights Activists

This study explored personal experiences of animal rights and environmental activists in New Zealand. The stories of participants provided insight into the challenges activists face in a country where the economy is heavily dependent on animal agriculture. A qualitative methodology was utilised and several major themes emerged: (1) emotional and psychological experiences, (2) group membership, (3) characteristics of activism and liberation, (4) the law and its agents, and (5) challenge to society. Participants of the study represent a group of individuals engaged in acts of altruistic offending triggered by exposure to the suffering of non-human animals. Their moral philosophy and conscience overrode all considerations for legal repercussions, and through their activism they not only challenged the status quo, but also called upon non-activist members of society to make meaningful contributions to the world around them

Retardation of cochlear maturation and impaired hair cell function caused by deletion of all known thyroid hormone receptors

The deafness caused by early onset hypothyroidism indicates that thyroid hormone is essential for the development of hearing. We investigated the underlying roles of the TRa1 and TRß thyroid hormone receptors in the auditory system using receptor-deficient mice. TRa1 and TRß, which act as hormone-activated transcription factors, are encoded by the Thra and Thrb genes, respectively, and both are expressed in the developing cochlea. TRß is required for hearing because TRß-deficient (Thrb tm1/tm1) mice have a defective auditory-evoked brainstem response and retarded expression of a potassium current (I K,f) in the cochlear inner hair cells. Here, we show that although TRa1 is individually dispensable, TRa1 and TRß synergistically control an extended array of functions in postnatal cochlear development. Compared with Thrb tm1/tm1 mice, the deletion of all TRs inThra tm1/tm1 Thrb tm1/tm1mice produces exacerbated and novel phenotypes, including delayed differentiation of the sensory epithelium, malformation of the tectorial membrane, impairment of electromechanical transduction in outer hair cells, and a low endocochlear potential. The induction ofI K,f in inner hair cells was not markedly more retarded than in Thrb tm1/tm1mice, suggesting that this feature of hair cell maturation is primarily TRß-dependent. These results indicate that distinct pathways mediated by TRß alone or by TRß and TRa1 together facilitate control over an extended range of functions during the maturation of the cochlea

The ankle-link antigen: an eiptope sensitive to calcium chelation associated with hair bundles and the calycal processes of photoreceptors.

A monoclonal antibody, mAb E40, that specifically recognizes hair cells and photoreceptors was derived from a mouse immunized with a membrane fraction prepared from the sensory maculae of the chick inner ear. In the mature chick inner ear, punctate labeling is observed along each stereocilium, but staining is mostly concentrated around the basal end of the sensory hair bundles, where it is closely associated with surface specializations known as ankle links. The epitope recognized by mAb E40 is therefore referred to as the ankle-link antigen (ALA). During early embryogenesis, the ALA is initially distributed evenly over the surface of the hair bundle. As development proceeds, it becomes more restricted to the base of the hair bundle, although a spot of the ALA remains associated with the bundle tip until just before hatching. In the eye, mAb E40 stains the calycal processes of photoreceptors. When maculae and retinae are treated with the calcium chelator BAPTA at room temperature, the ALA disappears. BAPTA-induced loss of the ALA from the hair-bundle surface is substantially reduced by lowering the temperature to 2 degrees C. The ALA and ankle links reappear on the hair-bundle surface when cells are cultured for 20 hr after BAPTA treatment. BAPTA sensitivity and recovery after BAPTA-induced loss are properties similar to those described for the tip link, a surface structure thought to gate the mechanotransducer channel. However, unlike the tip link, the ALA and ankle links are sensitive to subtilisin treatment. The results define a new component of the hair-bundle surface, with properties both common to and distinct from those of the tip link