DC potentials in different cells of the stria vascularis measured in vitro

Fine glass microelectrodes were inserted into the stria vascularis (SV) from the endolymphatic side under conditions in vitro. DC potentials were recorded in individual cells of the SV. The electrophoretic dye marking technique was used for identification of cells with different DC potentials. Penetration of the luminal membrane of marginal cells (MC) was accompanied by an unstable transient negative potential of about -15 mV. When the electrode penetrated further into the MC, a positive potential of about +10 mV was recorded. In the basal cells (BC) of the SV a negative DC potential of approx. -45 mV was always measured. Addition of ouabain into the perfusion solution (10(-3) mol/l) abolished the positive potential of MC, whereas the negative potential of BC decreased to approximately one half of the original value. A higher positive potential (+17 mV) was found in the MC of animals which were exposed to noise twenty days before the recording. Results are discussed in the light of the knowledge about localization of different transporting systems within the SV cells.     

Profiles of resting potentials across the stria vascularis in kanamycin-deafened guinea pigs

The resting potentials of the marginal cells in the stria vascularis of the guinea pig were determined from changes in the combined electrode-tissue resistance of the electrode. The resistance of the electrode was 45.5±16.0 MΩ (n=20) before penetration of the stria vascularis and 46.7±17.3 MΩ (n=20) after penetration. The resistance drops across the luminal membrane of the marginal cells were 46.0±22.6 MΩ (n=12) in kanamycin-deafened guineal pigs and 54.5±33.1 MΩ (n=9) in normal guinea pigs. The endocochlear potential (EP) and resting potentials in the marginal cells were 90.1±6.0 mV (n=14) and 70.4±11.3 mV (n=14) in kanamycin-deafened guinea pigs and 84.8±5.1 mV (n=29) and 74.7±11.7 mV (n=29) in normal guinea pigs. The resting potentials in the marginal cells decreased gradually and were approximately 0 mV around 20 min after anoxia in both kanamycin-deafened and normal guinea pigs. These changes were comparable to those of EP in kanamycin-deafened guinea pigs during anoxia. The mechanism of the EP in kanamycin-deafened guinea pigs is discussed.     

Adenine nucleotides of the stria vascularis

Summary The levels of the adenine nucleotides ATP, ADP, and AMP in the stria vascularis were measured under normal conditions, and following various durations of ischemia. The concentrations of these compounds were used for the calculation of the adenylate energy charge, the energy status and the phosphorylation state of the stria. Following 10 min of ischemia the adenylate energy charge had decreased three fold, the energy status seven fold and the phosphorylation state 14 fold. To study the potential for recovery of strial function following various brief and prolonged ischemic intervals, a method for the perfusion of the ear via the anterior inferior cerebellar artery was developed. For various reasons it was found advantageous to use artificial blood as perfusate, relying upon fluorocarbons as oxygen carriers. The endolymphatic potential was used as electrical indicator of strial function. Recovery of the endolymphatic potential following brief periods of ischemia was paralleled by a corresponding increase of the ATP levels and a drastic decrease of the AMP levels of the stria vascularis. Preliminary results on the effects of substrate-free perfusion are presented.Supported by NIH grant NS 06575, and NSF grant BNS 77-16842     

Scanning ultrastructure of the stria vascularis

In the eleven years since the introduction of the first commercially available scanning electron microscope, a number of otologic studies have made use of this new technique. However, the literature contains only one report of scanning ultrastructure of the endolymphatic surface of the stria vascularis. We have studied the stria vascularis with a technique allowing examination of the mid-modiolar cut surface as well as the endolymphatic surface.     

Scanning ultrastructure of the stria vascularis

Summary In the eleven years since the introduction of the first commercially available scanning electron microscope, a number of otologic studies have made use of this new technique. However, the literature contains only one report of scanning ultrastructure of the endolymphatic surface of the stria vascularis. We have studied the stria vascularis with a technique allowing examination of the mid-modiolar cut surface as well as the endolymphatic surface.Abbreviations used Figure legends — 1SP lower spiral prominence - uSP upper spiral prominence - E endolymphatic - CS cut surface - c capillaries - S sinus - SL spiral ligament Text abbreviations SEM scanning electron microscope (microscopy) - TEM transmission electron microscope - Å angstrom     

Passive transport of the stria vascularis

Summary The transport and diffusion of the stria vascularis were studied by injection of ferritin and horseradish peroxidase into the endolymphatic space of the guinea pig. The observation was made by electron microscopy under the conditions with or without the apposition of NaCl crystals on the round window membrane.In the cochlea with no NaCl crystals on the round window membrane, ferritin or HRP was found only in the apical cytoplasm of the marginal cells. No trace of particles was seen in the intermediate, basal, endothelial cells and intercellular spaces in the stria vascularis.The ferritin passed through the marginal cells, intercellular spaces, basement membrane and capillary walls by vesicular transport under the condition of change of osmotic pressure by apposition of NaCl crystals on the round window membrane.HRP labeled the marginal cells but was not seen in the intermediate, basal, endothelial cells and intercellular spaces.The functional significance of these findings were discussed.

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Zusammenfassung Transport und Diffusion der Stria vascularis wurden durch Injektion von Ferritin und Meerrettich-peroxidase (MRP) in den Endolymphraum des Meerschweinchens untersucht und die Veränderungen elektronenmikroskopisch dargestellt. Einem Teil der Tiere wurden Kochsalzkristalle auf die Membran des runden Fensters appliziert. Bei den Schnecken ohne Kochsalzkristalle am runden Fenster wurde Ferritin oder MRP nur im apikalen Teil des Cytoplasmas der Marginalzellen gefunden. Die intermediären, die basalen und die Endothelzellen sowie die intercellulären Räume der Stria vascularis waren frei.Bei Änderung des osmotischen Druckes nach Einlage von Kochsalzkristallen auf die runde Fenstermembran passiert Ferritin die Marginalzellen, die intercellulären Räume, die Basalmembran und die Capillarwände mittels vesiculären Transports. MRP fand sich in den Marginalzellen, nicht aber in den intermediären, basalen und endothelialen Zellen und den intercellulären Räumen. Die funktionelle Bedeutung dieser Befunde wird diskutiert.

     

Development of the stria vascularis in the rat

The rat is an altricial animal and is thus a useful model for the study of auditory development. The endocochlear potential (EP) undergoes a rapid increase in magnitude from the end of the 1st week to the beginning of the 3rd postnatal week. The purpose of this study was to examine the ultrastructure of the developing stria vascularis in the rat pup in order to correlate functional changes with structural alterations. Rat pups of various ages underwent EP measurement under Rompun anesthesia. The cochleas were rapidly removed under deep pentobarbital anesthesia. The tissues were fixed in 2.5% glutaraldehyde and postfixed in 1.5% osmium tetroxide. Thin sections were viewed and photographed using a Hitachi H7000 transmission electron microscope. A series of distinct developmental changes were observed. Intermediate and basal cells became more distinct from one another, and basal cells became more elongated. Marginal cells underwent progressive development of basolateral infoldings. These cytologic changes may signal the development of ion transport mechanisms necessary for EP development.     

Development of the human stria vascularis.

Fifteen human fetal cochleas were investigated by light microscopy and transmission electron microscopy in order to observe the development of the stria vascularis. The earliest signs of strial cell differentiation take place during the 11th week of gestation. Subsequently, the first stages of the stria vascularis development occur quickly. At week 14 the three types of cells, namely, marginal, intermediate and basal cells are discernable. Moreover at this stage, signs of specific activity are already present. The adult-like appearance of the stria vascularis is reached by week 21 but its maturation is completed only during the last trimester of pregnancy. This is in good agreement both with the development of the organ of Corti structures and with the maturation of the human auditory function.     

Quantitative assessment of the rat stria vascularis

Stria vascularis tissues from standardized regions in the basal, middle and apical turns of the rat cochlear duct were assessed quantitatively. Strial width, number of marginal cells across the strial width, radial area, as well as the volume density of the different components of the stria vascularis were determined for each standardized region. Strial width, number of marginal cells across the strial width and the radial area were greatest in the basal region and least in the apical region of the cochlea. The volume density of intermediate cells and capillary space was statistically unchanged in the three examined regions of the stria vascularis. However, the volume density of marginal cells and that of basal cells were different between regions. The volume density of marginal cells was highest in the basal turn while the volume density of basal cells was greatest in the apical turn. An objective assessment of the response of the stria vascularis to environmental conditions can be made by kant of its cellular architecture, providing a means to compare the effects of various agents between animal models used to study human inner ear dysfunction.     

Response of the stria vascularis to corticosteroids

Structural changes in the cellular architecture of the stria vascularis in adrenalectomized rats were quantitated by stereological methods after the administration of either aldosterone or dexamethasone. The volume densities of the differing strial components from steroid-administered animals were determined to approximate those of sham-adrenalectomized animals in general. The increased volume density of intercellular space as was observed following adrenalectomy, however, was only restored after the administration of aldosterone. These data correlate a recovery of the cellular architecture of the stria vascularis in adrenalectomized animals with the restoration of endogenous levels of adrenal steroids. These findings provide further information with regard to the effects of varying serum levels of corticosteroids on inner ear morphology.     

Adenine nucleotides of the stria vascularis.

The levels of the adenine nucleotides ATP, ADP, and AMP in the stria vascularis were measured under normal conditions, and following various durations of ischemia. The concentrations of these compounds were used for the calculation of the adenylate energy charge, the energy status and the phosphorylation state of the stria. Following 10 min of ischemia the adenylate energy charge had decreased three fold, the energy status seven fold and the phosphorylation state 14 fold. To study the potential for recovery of strial function following various brief and prolonged ischemic intervals, a method for the perfusion of the ear via the anterior inferior cerebellar artery was developed. For various reasons it was found advantageous to use "artifical blood" as perfusate, relying upon fluorocarbons as oxygen carriers. The endolymphatic potential was used as electrical indicator of strial function. Recovery of the endolymphatic potential following brief periods of ischemia was paralleled by a corresponding increase of the ATP levels and a drastic decrease of the AMP levels of the stria vascularis. Preliminary results on the effects of substrate-free perfusion are presented.     

Freeze fracturing of the human stria vascularis

The stria vascularis is an important functional element in the mammalian cochlea. This special tissue is considered to be the source of the endocochlear potential and thus the driving force for the production of a receptor response to the auditory stimulus. In order to maintain its function, the stria vascularis needs to be separated from the endolymphatic space by a tight seal. This seal is comprised of tight junctions in the marginal cell layer. The junctional arrangement in the stria vascularis is described, utilizing the freeze-fracturing technique which allows the visualization of large expansions of plasma membrane. The marginal cells are generally separated by tight junctions of the moderately tight to tight type. In places, however, even so-called leaky junctions with only a few sealing strands are present. Whereas the intermediate cell layer seems to lack tight junctions, the basal cells are connected by extensive tight junctions more or less covering the entire cell. These junctions seem to form an extremely tight barrier against the spiral ligament. Gap junctions are also present in the tissue. Intermediate cells as well as the basal cells are coupled by gap junctions. In the basal cell layer, gap junctional elements may also be found inside the large tight junctions comprising so-called mixed junctions.     

Freeze-etch visualizations of some elements in the stria vascularis

A quick-freeze, deep-etch study was made to observe the three-dimensional aspect of both membranes and organelles in the stria vascularis of the guinea pig. Our replicas showed various membrane characteristics of the vesicles, e.g., a clathrin basket on the inner surface membrane, aggregation of membrane particles on the protoplasmic fracture face, and regularly arranged surface protuberances on the outer surface membrane. In the cell processes of marginal cells, there were not only bundles of microtubules but also another type of fibril. Many gap junctions were observed between marginal, intermediate, and basal cells. In some replicas, the inner surfaces of the gap junctions were covered with particle protrusions with some ordered arrangements. The functional roles of these elements in the stria vascularis are discussed.     

Marginal cells of the stria vascularis in vitro]

Explants of stria vascularis and spiral ligament of the guinea pig cochlea were cultivated and after 2 days fibroblast-like cells were found growing around the explant. Marginal cells advanced at 15 microns/day to the border of the explant, and after 2 weeks they proliferated on top of a thin layer of fibroblast-like cells outside the explant, replacing several layers of fibroblast-like cells. Tight junctions and interdigitations of the lateral membranes were found between all neighbouring marginal cells. Their apical surface was covered by microvillus-like membrane extensions. The basal membrane of the new marginal cells did not interdigitate with the underlying membranes of fibroblast-like cells; there was always a gap between the two cell types. The results demonstrate that marginal cells of the stria vascularis are capable of repairing damage to the epithelium, such as may be caused by endolymphatic hydrops, even if the luminal side contains perilymph-like fluid. Furthermore, the cell culture allows living, clearly identified marginal cells to be studied in vivo.     

In vivo capillary diameters in the stria vascularis and spiral ligament of the guinea pig cochlea

Blood microvessels in the membraneous lateral wall of the cochlea were examined using intravital microscopic techniques. A video analysis system made serial diameter measurements at 1 micron intervals along the length of selected vessel segments during four experimental conditions. For each vessel segment, the serial measurements were statistically converted into a single diameter estimate, such that the flow resistance in a uniform vessel of this diameter would equal the resistance of the real non-uniform vessel. Nominal vessel diameters found (spiral ligament: 9-12 micron; stria vascularis: 12-16 micron) were nearly double those reported earlier in histological observations (Axelsson, 1968). During stimulation the largest diameter change seen was a 3.7% dilation (about 0.5 micron) in response to breathing 5% CO2 in oxygen. Theoretically, this change could reduce vascular fluid resistance by 16%, nearly enough to explain the observed flow increase of 20%. No diameter changes occurred for 5% CO2 in air despite a 50% flow increase, nor for air pressure pulses applied at the tympanic membrane. Round window electrical stimulation of 50 microA also produced dilation (less than 2.5%), but higher current levels were ineffective. In general, blood flow increases seen in this study could not adequately be attributed to the small lateral wall vessel diameter increases nor systemic causes, suggesting that lateral wall blood flow in these instances is dependent on control within the modiolus.