Recent and Planned Presentations by our group
Last updated November 25, 1996
Recent
Evidence for two components of endolymph volume regulation involving local and flow-based mechanismsA.N. Salt and J.E. DeMott
Association for Research in Otolaryngology, St. Petersberg Beach, Florida, February 1996.
Perilymph potassium increases during acute endolymph volume enlargement.
J.E. DeMott and A.N. Salt
Association for Research in Otolaryngology, St. Petersberg Beach, Florida, February 1996.
Endocochlear potential and cochlear fluid pH changes induced by respiratory CO2 manipulations
M. Suzuki and A. N. Salt
Association for Research in Otolaryngology, St. Petersberg Beach, Florida, February 1996.
Endolymph calcium homeostasis in pigmented and albino guinea pigs
S.S.Gill and A. N. Salt
Association for Research in Otolaryngology, St. Petersberg Beach, Florida, February 1996.
Endolymph volume regulation mechanisms revealed by microinjections into scala media..
Alec N. Salt and John E. DeMott
130th meeting of the Acoustical Society, St. Louis, November 1995.
Measurement of longitudinal fluid movements in the cochlea under normal and abnormal conditions.
Alec N. Salt
Symposium on "Intracranial and intralabyrinthine fluids - Basic aspects and clinical applications.", Seeheim, Germany, September 1995
Small longitudinal movements of endolymph are induced by volume disturbances.
Alec N. Salt and John E. DeMott
16th Danavox symposium on Meniere's disease, Kolding, Denmark, September 1995
Longitudinal movments and area changes of endolymph produced by osmotic dehydration.
J.E. DeMott and A.N. Salt
Association for Research in Otolaryngology, St. Petersberg Beach, Florida, February 1995.
Quantification of endolymphatic hydrops in the cochlea by magnetic resonance microscopy
Salt, A.N., Henson, M.H. Gewalt, S.L, Keating, A. W., DeMott, J.E. and Henson, O.W.
Association for Research in Otolaryngology, St. Petersberg Beach, Florida, February 1995.
Endolymph calcium differences between pigmented and albino guinea pigs.
S.S. Gill and A.N. Salt
Association for Research in Otolaryngology, St. Petersberg Beach, Florida, February 1995.
Endolymph Composition Changes in the Dominant Spotting (Wv) Mouse.
D. M. Miller and A.N. Salt
Association for Research in Otolaryngology, St. Petersberg Beach, Florida, February 1995.
Planned
Undetectable pressure increase during induction of acute endolymphatic hydrops by microinjection.J.E. DeMott and A.N. Salt
Association for Research in Otolaryngology, St. Petersberg Beach, Florida, February 1997.
Fixation-induced shrinkage of Reissner's membrane and its potential
influence on the assessment of endolymphatic volume
A.S. Brunschwig, A.N. Salt, J.E. DeMott, M.M. Henson and S.L. Gewalt
Association for Research in Otolaryngology, St. Petersberg Beach, Florida, February 1997.
Dependence of endolymph pH on perilymph bicarbonate concentration
Y.L. Ma, A.N. Salt and M. Suzuki
Association for Research in Otolaryngology, St. Petersberg Beach, Florida, February 1997.
Prior studies by have shown that injection of artificial endolymph into the cochlea at rates less than
60 nl/min for periods up to 15 min produced acute endolymphatic hydrops and /or basally-directed
flow of endolymph (Salt & DeMott ARO 19: #543, 1996). In the present study, microinjections into
the endolymphatic or perilymphatic spaces were performed while hydrostatic pressure was monitored
simultaneously in both compartments using micropressure instrumentation. Time averaging was performed
to reduce background pressure fluctuations due to respiration. During injections, virtually identical pressure
changes were recorded in both the endolymphatic and perilymphatic compartments. Digital subtraction of the
traces demonstrated no detectable change in pressure of endolymph with respect to perilymph during injection
into endolymph. It was estimated that less than 0.1 mm Hg pressure increase of endolymph with respect to
perilymph occurred while acute hydrops was produced. These results demonstrate that the terms endolymphatic
hydrops and endolymphatic hypertension are not synonymous. In the normal animal, the boundary membranes
of the endolymphatic system appear to be highly compliant so that pressure differences across the membranes
are extremely low. These results are in agreement with those of Takeuchi et al. (Ann. Otol. Rhinol. Laryngol.
100, 244, 1991) in which larger volumes of artificial endolymph were injected more rapidly than in the present
study. The predominantly basally-directed endolymph flow observed during endolymphatic injections in our
prior study indicates that a structure outside the cochlea, probably the saccule or endolymphatic sac, is even
more compliant than the structures bounding scala media in the cochlea.
Study supported by NIH/NIDCD DC01368
The quantification of endolymph volume by histological techniques or by magnetic
resonance (MR) microscopy could be influenced significantly by shrinkage of Reissner's
membrane during fixation. Since Reissner's membrane is anchored to bone at its medial
and lateral sides, shrinkage of a distended membrane would tend to pull it towards a
straight position. The goal of this study was to determine the amount of shrinkage of
Reissner's membrane induced by various fixation protocols and to establish a protocol
suitable for MR microscopy which minimizes soft tissue shrinkage. Fresh guinea pig
cochleae were dissected in an artificial perilymph, and fragments of Reissner's membrane
were isolated. Each individual specimen was observed in a petri dish using an inverted
microscope and its size documented. A selected fixative was then infused while the
specimen was observed and recorded on video tape. Size changes of the specimen were
quantified, usually over a 20 minute period. Heidenhain-Susa, a fixative widely used
for histological study of cochleae, caused substantial shrinkage of Reissner's membrane
during the first few minutes. The mean decrease in specimen length was 15.1 % ± 1.1%
(SE; n=8). It can be shown that the distension of Reissner's membrane in mild to
moderate hydrops would be nullified by a 15% shrinkage of the tissue. Other fixation
protocols produced far less shrinkage. The protocol showing least shrinkage was 3.1%
glutaraldehyde in Hanks' buffered salt solution, which produced a mean length decrease
of 0.3% ± 1.5% (SE; n=4).
For MR microscopy, mercuric chloride (HgCl2), a component of Heidenhain-Susa, greatly
enhances the contrast of Reissner's membrane against the background of the cochlear
fluid spaces, which appear white in the MR image. We have found that Reissner's
membrane looks black in specimens fixed with HgCl2-containing solutions, whereas
it appears white in formalin-fixed specimens. However, the inclusion of HgCl2 in
fixation media results in greater shrinkage of Reissner's membrane, when used as
either a primary or secondary fixative. Protocols for MR microscopy are presently
being optimized to maximize contrast while minimizing Reissner's membrane shrinkage.
In conclusion, the degree of Reissner's membrane shrinkage shown here to be caused by
some fixatives could result in systematic errors in the quantification of endolymph
volume.
This study was supported by NIH grants numbered DC01368 (AS), DC00114 (MH) and
1P41RR05959 (SG)
In previous studies, we found endolymph pH to be highly sensitive to respiratory CO2 status. Hypercapnea decreased endolymph pH while hypocapnea elevated pH. When the perilymphatic scalae of the cochlea are perfused with CO2-equilibrated artificial perilymph, we consistently observe an elevation in endolymph pH. We have found that one factor which contributes significantly to perfusion-induced endolymph pH changes is the concentration of bicarbonate in the perfusion medium. Since bicarbonate provides the dominant pH buffering in the perfusate, solutions of different bicarbonate content, equilibrated to physiological CO2 levels (approximately 5 %), will vary in pH, with pH increasing as bicarbonate increases. When the cochlea was perfused with CO2-equilibrated high-bicarbonate medium (50 mM HCO3), which has a higher pH than normal (pH 7.65), endolymph pH was decreased by an average of 0.05 (n=4). Conversely, perfusion with a nominally HCO3-free solution, caused endolymph pH to rise by an average of 0.11 (n=6). Both high- and low-bicarbonate manipulations generally reduced the endocochlear potential and elevated cochlear action potential thresholds. The observation that the change in endolymph pH is opposite in direction to the change in perilymph pH suggests that mechanisms of bicarbonate or pH homeostasis are involved. Far smaller endolymph pH changes were measured when comparable perilymph pH changes were induced by perfusing HCO3-free, HEPES-buffered solutions of varying pH. This supports the view that the observed endolymph pH changes with HCO3-modified media may be related to perilymph bicarbonate homeostasis. One explanation of the observations is that when perilymph bicarbonate is high, the secretion of acid to neutralize the excess perilymph bicarbonate, would release additional CO2, which, by rapidly equilibrating across membranous boundaries, would produce an acidification of endolymph. Conversely, under low-HCO3 conditions, the generation of bicarbonate from the reaction of CO2 and water, catalyzed by carbonic anhydrase, would reduce CO2 levels, resulting in an alkalinization of endolymph. These studies demonstrate that in the cochlea there are strong interrelationships between CO2, bicarbonate and fluid pH, in which the homeostasis of bicarbonate plays a highly prominent role.
This study was supported by NIH/NIDCD, grant number DC01368
Cochlear fluid movements can be accurately quantified with the use of chemical flow markers. In our technique, we injected small amounts of ionic markers (hexafluoro- arsenate or tetramethylammonium) into one of the cochlear fluids of pigmented guinea pigs. Movements of the marker were monitored by one or more ion selective micro- electrodes, which were sensitive enough to detect micromolar, non-toxic, marker concentrations. Results were interpreted using mathematical models that combined the effects of marker movements by diffusion, by longitudinal flow and by clearance from the compartment. In endolymph of the normal cochlea, longitudinal flow rates were extremely small, with a mean rate that was basally-directed at 4 µm/min. This rate was not significantly different from zero. However, when endolymph volume was disturbed, by osmotic dehydration or by direct volume injection, longitudinal volume flows were induced which contributed to the homeostatic process. Osmotic dehydration of cochlear endolymph produced apically-directed flow at a rate that averaged 43 µm/min. Direct micro-injections into cochlear endolymph at rates from 20-60 nl/min produced basally- directed endolymph flow in the basal turn. A somewhat similar situation exists for cochlear perilymph. When marker injection and recording electrodes were meticulously sealed into the cochlea, longitudinal movements of perilymph were extremely low, averaging only 1.6 nl/min in an apical direction along the basal turn of scala tympani. In contrast, when the otic capsule was perforated, releasing cochlear fluid pressure, the rate of longitudinal flow averaged 1.0 µl/min, which was over 600 times the normal rate. This high rate was due to cerebrospinal fluid entering the cochlea through the cochlear aqueduct, which rapidly displaced perilymph. We can conclude that in the normal state, longitudinal fluid movements make virtually no contribution to the homeostasis of the major ions in cochlear endolymph and perilymph. However, longitudinal flows may be induced in some situations, the effects of which may far exceed the normal, local homeostatic processes. Since the chemical compositions of inner ear fluids are known to vary in different regions, it is likely that chemical disturbances of cochlear tissues will occur under these conditions. (Work supported by NIH grant DC01368).
Alec N. Salt and John E. DeMott, Department of Otolaryngology, Washington University School of Medicine, 517 South Euclid Avenue, St. Louis, MO, 63110, USA.
We have previously shown that in the normal cochlea, longitudinal endolymph movements are extremely small and make a negligible contribution to the homeostasis of endolymph. In the present study, longitudinal endolymph movements were measured in guinea pigs during experimentally-induced disturbances of endolymph volume. In separate studies, volume was either decreased by perilymphatic perfusion of hypertonic medium, or was increased by direct injection of an artificial endolymph into the second turn. Induced changes of endolymph flow and area were measured in the basal turn using an ionic marker technique. The method involved the placement of three electrodes into endolymph of the basal turn. One electrode contained a volume marker ion (tetramethylammonium or hexafluoroarsenate), from which a trace amount was continually injected into endolymph by iontophoresis. Two ion-selective electrodes, sensitive to the marker ion, were inserted into endolymph, one apical and one basal to the injection site. The measured time courses of volume marker concentration during experimental manipulations were interpreted with the aid of a computer model, which simulated ionic movements in the endolymph. Our results show that when endolymph volume is disturbed, longitudinal movements of endolymph are induced which contribute to the measured ionic changes. Osmotic dehydration results in an apically-directed endolymph movement, while injection into the second turn produced basally-directed endolymph movement. The movements appear to be mechanically-induced, resulting from distension of the boundary structures of endolymph. These findings help reconcile previous apparent contradictions in the literature and support the concept that some aspect of endolymph volume regulation occurs outside the cochlea, in structures such as the endolymphatic sac.
Alec N. Salt (Dept. of Otolaryngology, Washington University School of Medicine, 517 South Euclid Avenue, St. Louis, MO, 63110) and John E. DeMott (Dept. of Otolaryngology, Washington University School of Medicine)
Longitudinal movements of endolymph were quantified in vivo in the basal turn of the guinea pig cochlea during injections of artificial endolymph into the second turn at rates up to 70 nl/min. Flow measurements utilized the marker ion tetramethylammonium (TMA), which was iontophoretically injected into the basal turn. TMA dispersion was measured by two ion- selective microelectrodes, one placed 0.5 mm apical, and the other 0.5 mm basal to the TMA injection site. Longitudinal endolymph movements were calculated from the recorded TMA time courses . Prior to injections, endolymph flow rates were extremely low. Volume injection into the second turn induced basally-directed flows in the basal turn. The relationship between the measured rate and the injection rate was nonlinear, in which low injection rates produced proportionately less flow than higher rates. These data show that for injections at low rates, volume disturbances have primarily local effects, presumably generating local distension or being compensated by local homeostatic mechanisms. At higher rates, basally-directed endolymph flow becomes increasingly significant. These findings suggest that two, independent processes may be involved in the regulation of endolymph volume. [Work supported by NIH grant DC01368]
Undetectable pressure increase during induction of acute endolymphatic hydrops by microinjection.
J.E. DeMott and A.N. Salt
Department of Otolaryngology, Washington University School of Medicine,
St. Louis, MO, 63110, USA.
Fixation-induced shrinkage of Reissner's membrane and its potential influence
on the assessment of endolymphatic volume
A.S. Brunschwig, A.N. Salt, J.E. DeMott, M.M. Henson and S.L. Gewalt
Department of Otolaryngology, Washington University School of Medicine,
St. Louis, MO, 63110, USA.
Dependence of endolymph pH on perilymph bicarbonate concentration
Y.L. Ma, A.N. Salt and M. Suzuki
Department of Otolaryngology, Washington University School of Medicine,
St. Louis, MO, 63110, USA.
Measurement of longitudinal fluid movements in the cochlea under normal and abnormal conditions.
Alec N. Salt
Department of Otolaryngology, Washington University School of Medicine,
St. Louis, MO, 63110, USA.
key words : cochlea, endolymph, perilymph, endolymphatic hydrops, fistula
.
Small longitudinal movements of endolymph are induced by volume disturbances
(Work supported by NIH grant DC01368)
Endolymph volume regulation mechanisms revealed by microinjections into scala media.
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