Observations on the stria vascularis of the guinea pig cochlea and the changes resulting from the administration of the diuretic furosemide.

An electron microscope examination of the stria vascularis of guinea pigs and the structural changes occurring after administration of furosemide has been made. The use of ruthenium-red, which stains cell coats, has shown that the atria is impermeable to the passive diffusion of material from both the endolymph and the spiral ligament. The first changes after administration of furosemide are observed 4 hours after injection; small spaces develop between the cells. After 5--6 hours the spaces have enlarged but the tight junctions between marginal cells remain intact. The intermediate cells show signs of atrophy. In severely affected animals the intermediate cells have so shrunk that the spaces are very large and the marginal cells clearly resolved. These latter cells show no structural damage. In this severely affected tissue it is seen that the blood vessels are entirely surrounded by marginal cell extensions. It is suggested that the intermediate cells may have a regulatory function. The damage appears to be recoverable, since 6 hours after injection the spaces have decreased in size although some signs of intermediate cell atrophy are still apparent.     

Cell membrane alterations in the stria vascularis of the guinea pig after ethacrynic acid treatment studied by freeze-fracture.

A freeze-fracture examination of the stria vascularis during the first 2 h after injection of ethacrynic acid was performed. This showed a re-distribution of the particles on the membrane fracture faces of both marginal and intermediate cells. As oedematous spaces developed, particle-poor, vesicle-like structures were found associated with both cell types. The tight junctions at the apices of the marginal cells and around basal cells were unaffected.     

Ultrastructure of the stria vascularis and Reissner's membrane in experimental hydrops

A time-sequence study was made of the early ultrastructural changes of the stria vascularis and Reissner's membrane in the guinea pig after obliteration of the endolymphatic sac and duct. Pathological alterations of both the stria vascularis and Reissner's membrane were found to start in the apex of the cochlea. The morphological changes of the stria vascularis were characterized by an increase of vesicles in the marginal cells and by intercellular edema, followed by vacuolization and atrophy of marginal and intermediate cells. In Reissner's membrane extensive gaps in the mesothelial cell layer were observed together with intracellular pathology of the epithelial cells. The significance of these ultrastructural changes in the stria vascularis and Reissner's membrane with regard to the pathophysiology of the endolymphatic hydrops is discussed.     

Analysis of structural changes in the stria vascularis following chronic gentamicin treatment

Changes in the stria vascularis following chronic gentamicin treatment have been examined using quantitative methods. Albino guinea pigs were given gentamicin at 100 mg.kg-1.day-1 subcutaneously for 10 days. Comparisons were made of strial tissue from the treated animals sacrificed either 1 h or 4 weeks following the last injection with that from saline-injected controls. Strial width (spiral prominence to Reissner's membrane) and marginal cell (MC) number across the stria were determined from scanning electron micrographs. Strial thickness (endolymphatic surface to spiral ligament) and the volume fractions of the strial components (MCs, intermediate cells (ICs), basal cells (BCs) and capillaries) were derived from thin sections. Qualitative changes to both MCs and ICs were apparent 1 h after the last injection. At four weeks post-treatment, there was a small, but statistically significant, decrease in the number of marginal cells and a highly significant decrease in strial thickness. This was almost entirely due to a highly significant decrease in the volume fraction of MCs (i.e. shrinkage). The volume fraction of ICs was increased but this could be accounted for by MC shrinkage; after allowing for the reduction in strial thickness, the volume occupied by ICs was unchanged. Thus, following chronic gentamicin treatment, the stria is affected but significant progressive and permanent structural effects are confined to the marginal cells.     

The time course of the strial changes produced by intravenous furosemide

The ultrastructural abnormalities produced in the stria vascularis by intravenous furosemide (80 mg/kg) were investigated in 14 guinea pigs. The changes consisted of marginal cell swelling, shrinkage of the intermediate cells and enlargement of the intercellular spaces, as described in other intoxications. The cytological derangements (including characteristics dilatation of the Golgi membranes) differed in detail from those arising after a comparable dose of ethacrynic acid. The morphological alterations were already present at 2 min, were maximal at 10 min, recovered only slowly at first and had not disappeared entirely at 180 min. For comparison, the fall in the endocochlear potential had a latent period of 20 s and was greatest at 2.3 min; its recovery was rapid initially but also incomplete at 180 min. Thus, no gross discrepancy in the time courses occurred, even if the correlation was imperfect. That reported previously must be due, therefore, to the much longer delays found following intraperitoneal administration.     

Cell membrane alterations in the stria vascularis of the guinea pig after ethacrynic acid treatment studied by freeze-fracture

Summary A freeze-fracture examination of the stria vascularis during the first 2 h after injection of ethacrynic acid was performed. This showed a re-distribution of the particles on the membrane fracture faces of both marginal and intermediate cells. As oedematous spaces developed, particle-poor, vesicle-like structures were found associated with both cell types. The tight junctions at the apices of the marginal cells and around basal cells were unaffected.This work was supported by the Wellcome Trust in the form of a Research Fellowship     

速尿对离体培养豚鼠血管纹毒性作用的观察

目的：观察速尿对离体培养的豚鼠血管纹组织的影响，探讨速尿耳毒性的作用机制。方法：20只花色豚鼠随机分成二组：速尿组（n=16），正常对照组（n=4）。应用组织块培养的方法，将血管纹组织培养24小时，随即对不同的实验组分别应用不同终浓度的的速尿（60、300、600、1250、2500μg/ml），分别继续培养30分钟和90分钟，观察培养的血管纹组织学结构。结果：速尿组在组织学方面均未出现血管纹水肿、缘细胞胞浆肿胀、细胞间隙扩大和中间细胞皱缩等病理改变，与对照组相比较血管纹结构无显著性差异。结论：速尿对体外培养的豚鼠血管纹组织无明显诱导水肿的作用，提示速尿耳毒性的产生，可能是间接的作用机制。     

Degeneration of the stria vascularis during development in melanocyte-deficient mutant rats (Ws/Ws rats)

Developmental changes in the stria vascularis of white spotting (Ws) rats were examined by scanning and transmission electron microscopes and by diaminobenzidine-staining techniques. The stria of Ws/Ws homozygote rats was found to have both pigmented and non-pigmented portions. While the pigmented portions possessed intermediate cells in the same manner as the stria of wild +/+ rats, the non-pigmented portions lacked the cells. Examination at 1, 2, 3, 4, 6, 8 and 14 weeks after birth revealed a progressive degeneration in the marginal cells and strial capillaries in the non-pigmented portions. At 1 week, no significant differences were seen in the marginal cells of any of the rats examined. At 2 weeks, the basolateral infoldings of the marginal cells were seen to be well developed and adult-like in the pigmented portions of Ws/Ws rats and in +/+ rats. In the non-pigmented portions, the basolateral infoldings of the marginal cells appeared well developed; however, vacant spaces were seen around the basolateral infoldings. At 3 weeks, the basolateral infoldings of the marginal cells in the non-pigmented portions had become more atrophic, and the empty spaces around the basolateral infoldings had enlarged. Also, the marginal cells themselves had become flatter or thinner. These findings became more prominent at 4 weeks and 6 weeks. At 8 weeks and 14 weeks after birth, the marginal cells appeared markedly flat, and no basolateral infoldings were seen in the non-pigmented portions. Pigmented portions of the stria in Ws/Ws rats, on the other hand, showed normal development throughout this period. A DAB-staining examination of the stria capillary net in Ws/Ws rats showed it to be well developed at 3 weeks in both pigmented and non-pigmented portions. At 8 weeks, a thickening of the capillary basement membrane was apparent. The above findings lead the authors to believe that intermediate cells play an important role in the development and maintenance of marginal cells and the strial capillary system.     

The time-course of furosemide-induced strial changes in guinea pigs after pretreatment with daltroban

It was shown previously (Ernst et al., 1989) that pretreatment of guinea pigs with a thromboxane (TX) receptor antagonist attenuates the decline of the endocochlear potential (EP) induced by furosemide. The present paper is aimed at investigating a possible correlation between the electrophysiological data and ultrastructural changes of the stria vascularis by electron microscopy. The dosages of 40, 60, and 80 mg/kg furosemide were injected after the pretreatment with the TX receptor antagonist daltroban and compared to controls which were injected with furosemide only. It was found at all furosemide concentrations that the strial changes 10 min after injection were nearly unchanged against controls. 30 min after furosemide injection, the most pronounced changes were seen when pretreating the animals: a clear reduction of the marginal cell swelling and edema in general were observed at 40 and 60 mg/kg furosemide. The guinea pigs injected with 80 mg/kg furosemide after pretreatment displayed nearly the same changes as controls.     

Effects of furosemide on endocochlear potentials, auditory action potentials and summating potentials and the changes of inner ear pathology]

Guinea pigs were injected with furosemide 50 mg/kg (group A) and 25mg/kg (group B). Two minutes after injection, EP of group A decreased to -13.9mv while that of group B decreased to +65 mv. Also, AP of group A disappeared, and recovered at 8.5 mins. while AP amplitude of group B decreased to 78%. The SP value of group A changed from -14.5mv to +23.4mv 1 min after injection and returned to negative polarity in 12 min. Edema of stria vascularis was observed under light microscope. Transmission electron microscope showed edema between marginal cells and intermedia cells, cytoplasm of the marginal cell protruded to the cochlear duct, and cell membrane of outer hair cell folded. The finding of this study illustrates that furosemide inhibits the transportation of the active ions of cochlear duct tissue resulting in decrease of EP and alters the function of hair cells causing the change of AP amplitude. -SP depends on the ion transportation, the polarity can be inversed while large dosage of furosemide was used.     

GJB6基因敲除小鼠耳蜗血管纹超微结构观察

目的观察缝隙连接蛋白30（connexin，Cx30）基因敲除纯合子Cx30（-/-）小鼠（P3、P7、P10、P30、P60、P90）耳蜗血管纹（stria vascularis，SV）超微结构的发育情况，进一步探讨GJB6基因突变导致耳聋的机制。方法选取Cx30（-/-）小鼠P3、P7、P10、P30、P60、P90等6个发育阶段的小鼠，用透射电镜观察耳蜗血管纹超微结构变化情况。结果P3～P90耳蜗血管纹边缘细胞、中间细胞、基底细胞呈现一个由不成熟至成熟的正常发育过程，至P90时均未发现退化性表现或缺失改变。血管纹毛细血管随着小鼠日龄的增加，管腔逐渐变宽，逐渐成熟，高倍镜下见P3、P7、P10时血管内皮细胞结构无明显异常，P30时内皮细胞之间的纤维突起出现疏松，P60时内皮细胞之间出现较大的裂隙，P90时这种改变更加严重，内皮细胞之间出现扩大的细胞间隙。结论 GJB6基因缺陷本身并不影响耳蜗组织血管纹的形态发育和成熟，GJB6缺失引起小鼠出生后逐渐出现的血管纹内皮细胞超微结构的改变导致内皮细胞屏障破坏，可能引起耳蜗内电位缺失，从而造成听力的下降。     

Ultrastructural changes in the albino guinea pig cochlea at different survival times following cessation of 8-day cisplatin administration

OBJECTIVE: To investigate the effect of cisplatin administration on the ultrastructural morphology of the organ of Corti, stria vascularis and spiral ganglion. MATERIAL AND METHODS: Forty-eight guinea pigs were treated with cisplatin by daily i.p. injection at a dose of 1.5 mg/kg for eight consecutive days. Electrocochleography was performed at various survival times after the final application of cisplatin. The cochleae were subsequently examined using electron microscopy. RESULTS: Ultrastructural examination corroborated that, in cochlear turns showing complete loss of outer hair cells (OHCs) at the light microscopic level, OHCs were indeed missing and had been completely replaced by supporting cells. OHC loss, the number of affected OHCs and the degree of intracellular pathology in the OHCs in the 1-day, 1-week and 2-week survival groups were considerably higher than in the 4- and 8-week survival groups. All degenerated OHCs demonstrated ultrastructural features commonly associated with necrosis. No morphological signs of apoptosis were observed. Strial changes consisted of protrusion of the apical membrane of the marginal cells into the scala media, without any other histopathological changes. Intermediate-cell atrophy, apparent as translucent areas at the light microscopic level, consisted of an increase in intercellular space due to shrinkage of intermediate and marginal cells ultrastructurally. Ultrastructural examination of the spiral ganglion showed that vacuolation of the spiral ganglion cells, seen at the light microscopic level, was due to severe swelling of the mitochondria. CONCLUSION: The present results corroborate our previous light microscopic findings. However, the ultrastructural results do not allow a conclusion to be drawn concerning whether the observed recovery is due to the formation of new OHCs or to (self-)repair of damaged OHCs, although the latter is less likely.     

The endolymphatic sac and inner ear homeostasis. II: Effect of glycerol on the sensory end organs with or without colchicine pretreatment

The effects of glycerol and colchicine on the sensory end organs of the inner ear were investigated in mice. Glycerol alone induced a widening of the intercellular spaces lining vestibular dark and transitional cells as well as the marginal cells of the stria vascularis. This was noted within 30 min after the injection of glycerol and was normalized again within 4 h after the injection. Colchicine induced some morphological changes in the inner ear sensory cells, such as dissociation of Golgi complexes etc. These isolated glycerol or colchicine injections did not cause any signs of inner ear functional impairment. Treatment with glycerol following pretreatment with colchicine, however, induced marked inner ear dysfunction with impaired sense of balance and audition. The inner ear morphology revealed a combination of changes as compared with what was observed after isolated treatment with glycerol or colchicine i.e. edema of the stria vascularis, and vestibular dark and transitional cells as well as dissociation of Golgi complexes in the sensory cells. The cochlea showed moderate endolymphatic hydrops. These findings indicate that colchicine affects the inner ear fluid regulating mechanisms which may lead to severe functional derangement after additional glycerol treatment. It is conceivable that the present experiment may serve as a useful model for further studies on inner ear changes related to endolymphatic hydrops and Ménière's disease.     

Effect of furosemide on basal lamina anionic sites in guinea pig labyrinth

The authors studied the effects of acute furosemide administration on the basal lamina (BL) anionic sites in the stria vascularis, ampullar crista, and endolymphatic sac by using cationic polyethyleneimine (PEI). Furosemide was intravenously administered to albino guinea pigs with normal Preyer's reflexes. After 20 minutes, the bony labyrinth was removed and processed for histologic evaluation. Under a transmission electron microscope, a marked enlargement of the intercellular spaces was observed in the stria vascularis. The PEI distribution decreased significantly on the capillary BL in the stria vascularis and on the subepithelial BL in the sensory, transitional, and dark cell areas. However, no significant change was observed on the capillary BL or the subepithelial BL in the endolymphatic sac. These findings suggest that acute furosemide administration severely alters the distribution of the anionic sites in the strial capillary BL and in the subepithelial BL in the ampullar crista, but not in the capillary BL or the subepithelial BL of the endolymphatic sac.     

Ultrastructural changes in the albino guinea pig cochlea at different survival times following cessation of 8-day cisplatin administration

Objective To investigate the effect of cisplatin administration on the ultrastructural morphology of the organ of Corti, stria vascularis and spiral ganglion.

Material and Methods Forty-eight guinea pigs were treated with cisplatin by daily i.p. injection at a dose of 1.5 mg/kg for eight consecutive days. Electrocochleography was performed at various survival times after the final application of cisplatin. The cochleae were subsequently examined using electron microscopy.

Results Ultrastructural examination corroborated that, in cochlear turns showing complete loss of outer hair cells (OHCs) at the light microscopic level, OHCs were indeed missing and had been completely replaced by supporting cells. OHC loss, the number of affected OHCs and the degree of intracellular pathology in the OHCs in the 1-day, 1-week and 2-week survival groups were considerably higher than in the 4- and 8-week survival groups. All degenerated OHCs demonstrated ultrastructural features commonly associated with necrosis. No morphological signs of apoptosis were observed. Strial changes consisted of protrusion of the apical membrane of the marginal cells into the scala media, without any other histopathological changes. Intermediate-cell atrophy, apparent as translucent areas at the light microscopic level, consisted of an increase in intercellular space due to shrinkage of intermediate and marginal cells ultrastructurally. Ultrastructural examination of the spiral ganglion showed that vacuolation of the spiral ganglion cells, seen at the light microscopic level, was due to severe swelling of the mitochondria.

Conclusion The present results corroborate our previous light microscopic findings. However, the ultrastructural results do not allow a conclusion to be drawn concerning whether the observed recovery is due to the formation of new OHCs or to (self-)repair of damaged OHCs, although the latter is less likely.     

The cochlea of the spontaneously diabetic mouse

Summary We examined the cochleae of the spontaneously diabetic KK mice by using transmission electron microscopy. At the age of 3 months, the mice started to show evidence for glycosuria and hyperglycemia, and tissue sections showed beginning cochlear pathology. The pathological changes present were found to be limited to the stria vascularis: protrusions of marginal cells, swellings of intermediate cells and widening of intercellular spaces were the main findings seen. These changes progressed with age, but were not observed in age-matched nondiabetic 57BL/6 mice. The possible mechanism of diabetes causing cochlear pathology is discussed.     

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.     

The cochlea of the spontaneously diabetic mouse

Summary We used transmission electron microscopy to examine the cochleae of non-obese diabetic mice as animal models for human type I or non-insulin-dependent diabetes mellitus. Pathological changes were observed in the organ of Corti of the basal turn and in the stria vascularis of each turn. Major findings in the stria vascularis were protrusion or condensation of marginal cells, swelling of intermediate cells, and widening of the intercellular spaces. Principal findings in the organ of Corti involved degenerative changes of the outer and inner hair cells and replacement of hair cells by supporting cells. No prominent pathological changes were observed in the capillaries. The possible mechanism of diabetic involvement in cochlear pathology is discussed.     

The cochlea of the spontaneously diabetic mouse. I. Electron microscopic observation of KK mice

We examined the cochleae of the spontaneously diabetic KK mice by using transmission electron microscopy. At the age of 3 months, the mice started to show evidence for glycosuria and hyperglycemia, and tissue sections showed beginning cochlear pathology. The pathological changes present were found to be limited to the stria vascularis: protrusions of marginal cells, swellings of intermediate cells and widening of intercellular spaces were the main findings seen. These changes progressed with age, but were not observed in age-matched non-diabetic 57BL/6 mice. The possible mechanism of diabetes causing cochlear pathology is discussed.     

Effects of organic acids on the edema of the stria vascularis induced by furosemide.

Furosemide is a loop diuretic which is ototoxic. Investigations have shown the stria vascularis to be the target tissue of this ototoxic drug. The purpose of the present study was to investigate the effects of furosemide on the stria vascularis in chinchillas, in controls and in animals pretreated with the above organic acids. Control animals were injected with 0.5 ml alkalinized saline followed by furosemide IV 30 min later. Experimental animals received probenecid, penicillin or sodium salicylate IV. Thirty minutes later, furosemide was injected in the same dose as in the controls. The basal turn of the stria vascularis was rapidly removed at various times from 10 to 30 min after furosemide administration and processed for transmission electron microscopy. Control animals were found to have reversible edema of the stria vascularis. Experimental animals had variable findings. Those animals pretreated with penicillin had virtually no edema of the stria vascularis at any time. Salicylate and probenecid pretreated animals had significantly less edema from one to 10 min after furosemide injection, but more edema than controls at later times. These findings suggest a discrepancy between ultrastructural pathology and functional status of the cochlea in experimental animals pretreated with probenecid or sodium salicylate followed by furosemide. On the other hand, good structure function correlations were seen in controls and in experimental animals pretreated with penicillin.