Stiffness, compliance, elasticity and force generation of outer hair cells.

Isolated outer hair cells (OHCs) were partially sucked into especially designed cell capillaries allowing an experimental reconstitution of the cells' electroanatomy. The experimental approach separated the apical from the basolateral parts of the cells thus forming an artificial scala media and scala tympani. Resistance between both was 121 +/- 42 M omega. A sequence of negative and positive pressures was applied to the basal cell pole allowing "pulling" or "pushing" of the sensory cell investigated. The resulting length changes together with the known pressures allowed the estimation of an actual longitudinal compliance of 354 +/- 35 m/N. Following "pulling" OHCs tended to resume their initial shape after the force had ceased to be effective indicating elastic distortions. The calculated elasticity modulus of OHCs amounted to 6.1 +/- 3.4 kN/m2. From this data an actual longitudinal whole cell stiffness of OHCs of 3 x 10(-3) N/m was calculated. Ultrasound scanning of immobilized OHCs identified the cuticular plate (CP) and a central core between CP and basal cell pole as structures contributing to the cells' acoustic stiffness. Changes of the potential differences between the artificial scala media and scala tympani resulted in active length changes following the command voltage with a slope of delta 1/(1 x U) = 0.055 V-1. Assuming the validity of Hooke's law, the force generation associated with the active length changes can be calculated since the compliance is known.(ABSTRACT TRUNCATED AT 250 WORDS)     

Structure,pharmacology and function of GABAA receptors in cochlear outer hair cells

Summary There is evidence that the inhibitory neurotransmitter -aminobutyric acid (GABA) is released from some efferent olivocochlear nerve endings terminating at outer hair cells (OHCs). Using monoclonal antibodies against postsynaptic GABA_A receptor from bovine cerebral cortex we confirm the presence of GABA and benzodiazepine bindings sites of - and -subunits of GABA_A receptors at the basal pole of isolated OHCs. Whole-cell recording with viable OHCs revealed that the application of 10^–3–10^–8 M GABA to the cell surface was followed by a concentration-dependent hyperpolarization of the outer cell membrane. Hyperpolarization was increased in the presence of 2.5 × 10^–5 M chlorazepate, a benzodiazepine derivative. Electrophysiological effects caused by GABA alone or in combination with chlorazepate were specifically inhibited by 10^–6 M of the GABA-receptor antagonist picrotoxin. Moreover, 10^–5–10^–7 M GABA caused reversible slow elongation of the cylindrical hair cell body in OHCs examined. These neurotransmitter-induced motile responses were specifically blocked by 10^–4 M picrotoxin. The results suggest that a subpopulation of OHCs express - and -subunits of GABA_A receptors which both form a GABA/benzodiazepine-receptor complex at the basal pole of isolated OHCs. These receptors are thought to allow GABA which is released from efferent auditory nerve terminals to bind to the cell surface of OHCs, resulting in GABA_Areceptor activation. This probably gates a GABA_A-receptor-associated chloride channel in the postsynaptic OHC membrane, allowing hyperpolarization and elongation of the cell. Correspondence to: P.K. Plinkert     

Salicylate-induced morphological changes of isolated inner hair cells and outer hair cells from guinea-pig cochlea

Objectives

The aim of this study is to investigate the salicylate-induced morphological changes of cochlear inner hair cells (IHCs) and outer hair cells (OHCs).

Methods

IHCs and OHCs were acutely isolated from the guinea-pig cochlea. Cells were observed under the inverted microscope and 10 mmol/L sodium salicylate solutions or 0.01 mmol/L dexamethasone-plus-salicylate solutions were applied. The cell length or the ratio between the length and width was the indices of the morphological changes in cells.

Results

Isolated IHCs did not demonstrate any significant changes in sodium salicylate solutions in 20 min and in 40 min, whereas OHCs were shortened by the 10 mmol/L sodium salicylate to 83% in 20 min and 75% in 40 min. There were no significant differences between in the dexamethasone-plus-salicylate solutions and in the control solutions after 20 min and 40 min both in IHCs and OHCs.

Conclusions

Although salicylate affected the isolated OHCs from guinea-pig cochlea, IHCs were not changed morphologically by sodium salicylate applications. Dexamethasone inhibited the salicylate-induced morphological changes of OHCs.     

Stiffness,Compliance, Elasticity and Force Generation of Outer Hair Cells

Isolated outer hair cells (OHCs) were partially sucked into especially designed cell capillaries allowing an experimental reconstitution of the cells' electroanatomy. The experimental approach separated the apical from the basolateral parts of the cells thus forming an artificial scala media and scala tympani. Resistance between both was 121±42 MΩ. A sequence of negative and positive pressures was applied to the basal cell pole allowing “pulling” or “pushing” of the sensory cell investigated. The resulting length changes together with the known pressures allowed the estimation of an actual longitudinal compliance of 354 ± 35 m/N. Following “pulling” OHCs tended to resume their initial shape after the force had ceased to be effective indicating elastic distortions. The calculated elasticity modulus of OHCs amounted to 6.1 ±3.4 kN/m². From this data an actual longitudinal whole cell stiffness of OHCs of 3 X 10^?3 N/m was calculated. Ultrasound scanning of immobilized OHCs identified the cuticular plate (CP) and a central core between CP and basal cell pole as structures contributing to the cells' acoustic stiffness. Changes of the potential differences between the artificial scala media and scala tympani resulted in active length changes following the command voltage with a slope of Δ/(1XU)=0.055 V^?1. Assuming the validity of Hooke's law. the force generation associated with the active length changes can be calculated since the compliance is known. We calculated a force of 14 pN, an energy stored in the elastic cell of 3x10^?20 J. Assuming that 50–60 jointly innervated OHCs act as a motor unit, the resulting force amounts to 700 pN/nm. Interestingly, Gummer et al. (1981) measured a BM stiffness of 480 pN/nm. Thus, the force exerted by 50 OHCs seems to be sufficiently effective to induce an in situ movement.     

Active movements of the cuticular plate induce sensory hair motion in mammalian outer hair cells

It has been suggested that active motor capacities may explain the high sensitivity and sharp tuning of auditory transduction in the mammalian cochlea. Recently, slow and fast motile responses of isolated outer hair cells (OHCs) have been demonstrated. These consist of reversible length changes in the cylindrical cell body. Here we show that isolated OHCs are also capable of slow and fast movements of the cuticular plate (CP) which cause passive displacements of the stereocilia. The molecular mechanisms that underlie the slow movements appear to reside in the highly organized actomyosin cytoskeleton in the CP. Additional, fast movements of the CP were observed in OHCs which were exposed to alternating electrical fields. These rapid mechanical events followed the electrical stimulus cycle-by-cycle, and unlike the slow movements, did not depend on the actomyosin skeleton in the CP. The fast motility of the CP and the attached stereocilia suggest an additional active mechanism which may contribute to the sensitivity and the sharp tuning of the cochlea near threshold. By controlling the operating point of the reticular lamina and of the hair bundles, the slow CP and stereociliary movements may influence basilar membrane homeostasis and temporary threshold shift at high sound pressure levels.     

Acetylcholine, carbachol, and GABA induce no detectable change in the length of isolated outer hair cells

The mechanical and electrical properties of cochlear outer hair cells (OHCs) are suggested to modulate transduction by inner hair cells. These properties of OHCs are presumably regulated by efferent neurons which use several transmitters including acetylcholine (Ach) and gamma aminobutyric acid (GABA). Since it had been suggested that Ach causes isolated OHCs to shorten visibly, this study was designed to investigate whether GABA also alters the length of OHCs. OHCs were isolated from the guinea pig cochlea by mechanical dispersion after collagenase treatment. Cells were initially selected by strict morphological criteria. In addition they were only included in further studies if they attained a constant length during 10 min of superfusion with buffer solution. Neither GABA (20 microM: 100 microM), Ach (5 mM; 10 microM with 10 microM eserine) or carbachol (10 microM; 100 microM) altered OHC length when applied in iso-osmotic Hank's balanced salt solution (total number of cells tested, 72). If a change in length occurred it must have been smaller than 0.3 microns, our detection ability. In contrast, high potassium and variations in osmolarity changed hair cell length by 3-10% in agreement with other reports.     

豚鼠耳蜗单离外毛细胞的外向整流钾电流

目的 :观察豚鼠耳蜗单离外毛细胞 (OHC)的电生理特性 ,记录不同长度OHC的外向整流钾电流 ,分析区分外向整流钾电流所包含的通道电流成分 ,研究外向整流钾电流的动力学特征。方法 :采用酶消化法及机械分离OHC。运用全细胞膜片钳技术 ,在电压钳下记录K+ 通道电流。结果 :OHC的全细胞膜电容为 (30 .96±2 .79) pF(n =2 9) ,零电流电位 (30± 2 .1)mV(n =16 ) ,反转电位为 (- 5 1.6 7± 1.84 )mV(n =9)。不同长度OHC的外向整流钾电流存在系统差异 ,短OHC表现出大的钾电导 ,长OHC则相反。 10 0 μmol/L的氯化镉 (Cd Cl2 )抑制了OHC外向整流钾电流的最大电流幅度的 6 0 % ,且改变了电流的动力学特征 ,对峰电流的影响明显大于稳态电流 (P<0 .0 1,n =5 ) ;1mmol/L的四氨基吡啶 (4 AP)抑制了最大电流幅度的 4 3% ,没有改变电流的动力学特征。外向整流钾电流的激活符合Boltzmann方程 ,V1/ 2 =(- 11.0 7± 0 .2 6 )mV ,S =(6 .6 2± 1.74 )mV(n=13)。结论 :外向整流钾电流包含有钙离子激活的钾离子电流、外向延迟整流钾电流和A型电流     

A subpopulation of outer hair cells possessing GABA receptors with tonotopic organization

Summary The olivocochlear innervation has been postulated to regulate active mechanical processes in the mammalian cochlea. Histochemical studies led to the suggestion that a subpopulation of these efferent nerves, which predominantly terminate on outer hair cells (OHCs), are -aminobutyric acid (GABA)-ergic. By means of two monoclonal antibodies, we were able to visualize GABA_A-receptor immunoreactivity at the basal pole of isolated sensory cells. Both subunits of the GABA_A receptor, the - and -subunit, are known to form the transmembraneous GABA/ benzodiazepine-receptor complex and were present on OHCs. In addition, these inhibitory receptors were more numerous in the apical turns of the cochlea, indicating another criterion for distinguishing the apical from basal turns of the cochlea. These results support the concept that a subpopulation of axosomatic synapses at the basal pole of OHCs liberate the inhibitory neurotransmitter GABA into the synaptic cleft. Binding of the transmitter to these newly observed subsynaptic receptors is possibly followed by a change in OHC motility and a subsequent modulation of the movement of the basilar membrane.     

Potassium-induced slow motility is partially calcium-dependent in isolated outer hair cells

Low flow rate (0.6 microl/min) administration of high concentration potassium solutions (12.5, 25 and 37.5 mM) was tested for evoking slow-motility length changes in isolated, apical turn, guinea pig outer hair cells (OHCs) (length 65-80 microm; n = 38). Control OHCs (n = 16) showed a flow rate-dependent, reversible, longitudinal shortening of 0.5-3 microm during perfusion with normal saline. Potassium, an effective depolarizing agent for OHCs, induced a concentration-dependent cell shortening of 0.5-13 microm. These cell shape changes were reversible. The magnitude of shortening was significantly (p < 0.01) decreased in a calcium-free incubation medium (n = 8). The velocity of the shortening was 300 nm/s in the first 10 s after application of 37.5 mM K+ in a normal incubation medium and decreased to 100 nm/s during the next 10 s. Corresponding velocities in calcium-free solutions were 100 and 50 nm/s, respectively. K+-induced shortening velocities were not significantly different from control values after 30 s. It appears that K+-induced OHC shortening is sensitive to the calcium content of the incubation medium during the first 10 s. Higher flow rate (1.5 microl/min) administration of K+ makes the velocity and magnitude of slow motility of OHCs insensitive to the absence of calcium. These results highlight the fact that one of the critical technical points in fluid perfusion experiments with isolated OHCs is selecting a safe low flow rate of < 0.6 microl/min. At this perfusion rate, K+-induced OHC shortening is composed of both calcium-sensitive and -insensitive components.     

Linopirdine Blocks α9α10-Containing Nicotinic Cholinergic Receptors of Cochlear Hair Cells

Studies of the electrophysiological response to acetylcholine (ACh) in mammalian outer hair cells (OHCs) are hindered by the presence of a large potassium current, I _K,n, most likely mediated by channels containing the KCNQ4 subunit. Since I _K,n can be blocked by linopirdine, cholinergic effects might be better revealed in the presence of this compound. The aim of the present work was to study the effects of linopirdine on the ACh-evoked responses through 910-containing native and recombinant nicotinic cholinergic receptors. Responses to ACh were blocked by linopirdine in both OHCs and inner hair cells (IHCs) of rats at postnatal days 21–27 (OHCs) and 9–11 (IHCs). In addition, linopirdine blocked responses of recombinant 910 nicotinic cholinergic receptors (nAChRs) in a concentration-dependent manner with an IC₅₀ of 5.2 M. Block by linopirdine was readily reversible, voltage independent, and surmountable at high concentrations of ACh, thus suggestive of a competitive type of interaction with the receptor. The present results contribute to the pharmacological characterization of 910-containing nicotinic receptors and indicate that linopirdine should be used with caution when analyzing the cholinergic sensitivity of cochlear hair cells.     

Local mechanical properties of mouse outer hair cells: atomic force microscopic study

OBJECTIVES: Outer hair cells (OHCs) are capable of altering their cell length in response to changes in membrane potential. Due to this electromotility, OHCs probably subject the basilar membrane to force, resulting in cochlear amplification. To understand the mechanism of such amplification, knowledge of the mechanical properties of OHCs is required since the force produced by OHC electromotility is thought to depend on such properties. Various studies have been conducted to investigate the mechanical properties of guinea pig OHCs. With regard to mice, however, although various kinds of transgenic and knockout mice possess great potential as research models, the mechanical properties of mouse OHCs have not as yet been reported since the cells and/or tissues in the mouse hearing organ are relatively small and vulnerable to external stimuli, rendering sample preparation difficult. In this study, therefore, to establish indicators of the mechanical properties of OHCs in mice, such properties were measured by atomic force microscopy (AFM). METHODS: CBA/JNCrj strain male mice aged 10-12 weeks (25-30 g) were used. Cochleae were dissected out from the animal and both the basilar membrane and the organ of Corti were simultaneously unwrapped from the modiolus with forceps. Dissected coiled tissue was then incubated with an enzymatic digestion medium for 15 min. After digestion, OHCs were isolated by gently triturating the coiled tissue. Local mechanical properties of the OHCs were then measured by an indentation test using an AFM. RESULTS: Young's modulus and stiffness of the OHC in the apical turn of the mouse cochlea were 2.1+/-0.5 kPa and 4.4+/-1.2 mN/m, respectively. CONCLUSIONS: Young's modulus of the OHC in the apical turn of the cochlea in mice was roughly the same as that in the apical turn of the cochlea in guinea pigs; however, the stiffness of the former was about two times greater than that of the latter because the cell length of the former was shorter than that of the latter.     

Apoptotic Pattern of Cochlear Outer Hair Cells and Frequency-specific Hearing Threshold Shift in Noise-exposed BALB/c Mice

Objectives

Apoptosis of outer hair cell (OHC) can be identified through nuclear staining by specific nuclear changes. The change of filamentous actin (F-actin) is also involved in early cell death process. The study was designed to investigate OHC death along the whole length of the organ of Corti.

Methods

BALB/c hybrid mice were used in this study. The noise group was exposed to white noise of 120 dB SPL for 3 hr per day for 3 consecutive days. The tone burst auditory brainstem response (ABR) test was conducted and cochleas from each group were obtained for the immunostaining of FITC phalloidin for F-actin and propidium iodide (PI) for nuclei.

Results

ABR threshold of the noise group significantly increased after noise exposure (P<0.001). No threshold shift was found in the control group. Threshold shift of the noise group constantly increased from 4 to 16 kHz, but threshold shifts at 16 kHz and 32 kHz were similar. Patterns of OHC staining were subclassified as FITC+PI- cells, FITC+ PI+ cells, FITC-PI+ cells and missing cells. Proportion of normal live OHCs (FITC+PI-) rapidly decreased from the apex to the base. In the basal turn, FITC-PI+ cells and vacancy OHC (missing cells) were observed easily. Apoptotic and missing cells were most abundant at 60% of the whole length of the Corti organ.

Conclusion

We could subclassify morphologic changes in OHC death after noise exposure. Quantitative changes in OHCs along the whole Corti organ showed a plateau pattern similar to that of a frequency-specific threshold shift.     

Effect of corticosteroid on salicylate-induced morphological changes of isolated cochlear outer hair cells

Our previous studies showed that pretreatment with corticosteroids, which inhibits release of arachidonic acid (precursor of prostaglandins and leukotrienes), partially prevented salicylate-induced hearing loss in vivo. The purpose of this study was to determine the effect of pretreatment with corticosteroid (dexamethasone sodium phosphate) on isolated cochlear outer hair cells (OHCs) exposed to salicylate in vitro. Isolated OHCs from the chinchilla cochlea were exposed to salicylate with or without pretreatment with dexamethasone. Images were stored and analyzed on the Image program. The OHCs exposed to salicylate demonstrated a significant shortening in cell length. The OHCs exposed to salicylate after pretreatment with dexamethasone exhibited no significant change in cell length. We conclude that corticosteroid treatment of isolated OHCs is effective in blocking the morphological changes induced by salicylate. This study gives additional evidence that salicylate ototoxicity is mediated by alteration in the levels of arachidonic acid metabolites.     

Effect of aminoglycoside otic drops on isolated cochlear outer hair cells with and without liver extract activation

OBJECTIVE: To assess the ototoxicity of commercially available Gentacidin and TobraDex ear drops with and without liver extract activation using isolated cochlear outer hair cells (OHCs). MATERIAL AND METHODS: OHCs from adult chinchilla cochleae were exposed to standard bathing solution (SBS), liver extract alone and Gentacidin and TobraDex ear drops with and without liver extract. All experiments were performed at an osmolality of 305 +/-5 mOsm, at room temperature and for up to 60 min. OHC images were recorded using an inverted microscope and analyzed electronically. Time to cell death and changes in cell length were measured. RESULTS: The time to cell death and the percent change in cell length were significantly shorter in the Gentacidin+liver extract group than in the Gentacidin alone group (p < 0.05). The TobraDex+liver extract group showed a significantly decreased time to cell death compared to the SBS control group (p < 0.05). There were no significant differences in cell length or time to cell death between the TobraDex+liver extract group and the TobraDex alone group (p > 0.05). CONCLUSION: This study suggests that the cytotoxicity of aminoglycoside ear drops to isolated OHCs in vitro requires     

Effect of platelet activating factor with and without receptor antagonist (WEB2170) on morphology of isolated cochlear outer hair cells

Platelet activating factor (PAF), generated from biologically active phospholipids, has been implicated as a potent inflammatory mediator and has been shown to be involved in many pathological processes, especially in inflammation and allergy. It has been suspected that PAF may be one of the inflammatory mediators in middle ear effusion that can induce sensorineural hearing loss, as observed in chronic otitis media. The PAF receptor antagonist WEB2170 has been studied extensively, and its inhibitory effects against various PAF actions are well proven in otologic systems. The purpose of our study was to determine the effect of superfusion of PAF and WEB2170 on morphological changes in isolated cochlear outer hair cells (OHCs). Isolated OHCs from adult chinchilla cochleas were exposed to albumin-phosphate-buffered saline solution (1 mg/mL), WEB2170 (5 mg/30 mL), PAF (1 micromol/L), or both PAF (I micromol/L) and WEB2170 (5 mg/30 mL). All experiments were performed at an osmolality of 305 +/- 5 mOsm at room temperature for 30 minutes. The cells were observed with an inverted microscope; the images were stored and analyzed on the Image Pro-Plus program. The OHCs exposed to control albumin-phosphate-buffered saline solution or to WEB2170 did not show any significant change in cell shape or length. The cells exposed to 1 micromol/L of PAF showed ballooning and significant shortening of the mean cell length in 15 to 20 minutes. These morphological changes in OHCs can be prevented by pretreating OHCs with WEB2170. This study demonstrated that exposure to PAF causes morphological changes in isolated OHCs that can be prevented by the PAF receptor antagonist WEB2170.     

Inner hair cells isolation and morphological observation]

OBJECTIVE: To set up an effective technique for isolating the single inner hair cells (IHCs) and observe morphological features to distinguish IHCs from other hair cells (OHCs) in vitro. METHODS: Surface preparations contained rows of OHCs and IHCs were prepared from guinea pigs. The cluster and single IHCs were separated microsurgically combination with an enzymatic digestion. The clusters of IHCs were separated using two fine electrodes and transferred by a gentle suction using a glass micropipette. RESULTS: On the average, one guinea pig cochlear yielded approximately 30 to 50 viable solitary IHCs. The cell body of the IHCs was pear or flask shape. The nucleus was located in the central of the cell body, and the cytoplasm was filled with the structured and scattered rough granules. The stereocilia were seen in the most of IHCs (93 out of 98). The stereocilia were aligned, almost lineally or in a "C" shape, in one side on the surface of the cuticular plate. The tight neck and the angle between the cuticular plate and the axis of the cell were observed in some IHCs but not obvious in most cases, which might be due to the orientation of the cells. The length of isolated IHCs was ranged from 13 to 31 microns and with an average of (22.45 +/- 4.14) micron (mean +/- s, n = 98). The diameter of IHCs was ranged from 7 to 15 microns and with an average of (11.95 +/- 1.59) micron. The length of stereocilia was ranged from 2 to 7.5 microns and with an average of (5.21 +/- 1.00) micron. CONCLUSION: Unambiguous solitary IHCs were successfully harvested from guinea pig cochlea using a microsurgical technique. The tight neck and the angle between the cuticular plate and the axis of the IHCs have been considered as an important landmark to distinguish the IHCs from OHCs.     

Guinea pig vestibular type I hair cells can show reversible shortening.

Guinea pig isolated vestibular type I hair cells (VIHCs) were recently reported by our group to respond to high [KCl] solutions by an irreversible tilt of their neck region and sometimes by a sustained shortening and swelling. A possible osmotic contribution to these shape changes was investigated by substituting gluconate (G) for chloride in the test solution, so as to minimize water influx, and also by changing the osmotic pressure of the extracellular solution. For comparison, similar experiments were also undertaken on cochlear outer hair cells (OHCs). Utricular and ampullar type I hair cells were more difficult to isolate than OHCs and, like them, responded to an isotonic high [KCl] solution by a sustained shortening and widening, which were found to be reversible for most cells when rinsed with the control solution. In a high [KG] solution, all OHCs showed a shortening reversible in the test solution; among the VIHCs tested, two-thirds presented a slight sustained shortening without widening and a third showed a spontaneously reversible shortening, particularly at the neck level. VIHCs exposed to a high [N-methyl-D-glucamine chloride] solution, this impermeant cation replacing K+ for control, presented only a slight sustained shortening. In response to osmotic changes of the bathing medium, both VIHCs and OHCs showed a sustained shortening or elongation (the latter to a lesser degree) for hypo- and hyperosmotic solutions, respectively. The VIHCs and OHCs that presented a reversible shortening in a high [KG] solution widened concomitantly with their shortening, but to a smaller extent compared with what was observed in a high [KCl] solution, and this diameter increase was reversible in the test solution, unlike the widening observed in a hypotonic solution. These results show that a reversible shortening occurred for some VIHCs; they also indicate the involvement of two components in the KCl-induced response: one osmotic and another potassium-dependent.     

Modell einer Elektrode zur multiaxonalen Reizung des Nervus acusticus

Summary An electrode arrangement is described, which allows an isolated stimulation of 25–50 singular axons: By laser beams a small ceramic plate got 25–50 perforations through which glass isolated platin wires were so fixed that they end like a brush. Only the tips of the hairs are not isolated. The diameter of the electrodes is approximately 10 m, the length ranges between 0,5 and 1,5 mm. This electrode system is enough stabile to be pressed into a hard boiled egg without being damaged.     

Hypotonic swelling of salicylate-treated cochlear outer hair cells

The outer hair cell (OHC) is a hydrostat with a low hydraulic conductivity of Pf=3x10(-4) cm/s across the plasma membrane (PM) and subsurface cisterna that make up the OHC's lateral wall. The SSC is structurally and functionally a transport barrier in normal cells that is known to be disrupted by salicylate. The effect of sodium salicylate on Pf is determined from osmotic experiments in which isolated, control and salicylate-treated OHCs were exposed to hypotonic solutions in a constant flow chamber. The value of Pf=3.5+/-0.5x10(-4) cm/s (mean+/-s.e.m., n=34) for salicylate-treated OHCs was not significantly different from Pf=2.4+/-0.3x10(-4) cm/s (mean+/-s.e.m., n=31) for untreated OHCs (p=.3302). Thus Pf is determined by the PM and is unaffected by salicylate treatment. The ratio of longitudinal strain to radial strain epsilonz/epsilonc=-0.76 for salicylate-treated OHCs was significantly smaller (p=.0143) from -0.72 for untreated OHCs, and is also independent of the magnitude of the applied osmotic challenge. Salicylate-treated OHCs took longer to attain a steady-state volume which is larger than that for untreated OHCs and increased in volume by 8-15% prior to hypotonic perfusion unlike sodium alpha-ketoglutarate-treated OHCs. It is suggested that depolymerization of cytoskeletal proteins and/or glycogen may be responsible for the large volume increase in salicylate-treated OHCs as well as the different responses to different modes of application of the hypotonic solution.     

利诺吡啶对豚鼠耳蜗外毛细胞和支持细胞钾电流的影响

目的 观察KCNQ家族钾通道特异性阻滞剂利诺吡啶(linopirdine)对豚鼠耳蜗单离外毛细胞和Deiters细胞(支持细胞)总钾电流的影响，初步探讨KCNQ家族钾通道在耳蜗外毛细胞和Deiters细胞的分布。方法 运用膜片钳技术，在全细胞模式下记录正常细胞外液中8个外毛细胞和5个Deiters细胞的总钾电流，并观察100μmol／L利诺吡啶对外毛细胞和Deiters细胞总钾电流的影响。结果 在正常细胞外液中，单离外毛细胞可记录到四乙基二乙胺敏感的外向性钾电流和静息膜电位附近激活的内向性钾电流(the K^ current activated at negative potential，IKn)两种钾电流，而单离Deiters细胞中只记录到外向整流性钾电流。加入100μmol／L利诺吡啶后，外毛细胞中的四乙基二乙胺敏感的钾电流减小，IKn被完全抑制；而Deiters细胞中的外向整流性钾电流大小无变化。结论 KCNQ家族钾通道存在于豚鼠耳蜗外毛细胞，其介导的钾电流是四乙基二乙胺敏感的钾电流的组成部分，并构成全部的IKn；但KCNQ家族钾通道不存在于豚鼠耳蜗Deiters细胞。