Acta Oto-Laryngologica
ISSN: 0001-6489 (Print) 1651-2251 (Online) Journal homepage: http://www.tandfonline.com/loi/ioto20
Differences in Hair Bundles Associated with Type I and Type II Vestibular Hair Cells of the Guinea Pig Saccule Pascale Lapeyre, Anne Guilhaume & Yves Cazals To cite this article: Pascale Lapeyre, Anne Guilhaume & Yves Cazals (1992) Differences in Hair Bundles Associated with Type I and Type II Vestibular Hair Cells of the Guinea Pig Saccule, Acta Oto-Laryngologica, 112:4, 635-642 To link to this article: http://dx.doi.org/10.3109/00016489209137453
Published online: 08 Jul 2009.
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Date: 28 January 2016, At: 07:41
Acta Otolaryngol (Stockh) 1992; 112: 635-642
Differences in Hair Bundles Associated With Type I and Type I1 Vestibular Hair Cells of the Guinea Pig Saccule PASCALE LAPEYRE, ANNE GUILHAUME and YVES CAZALS From the Laboratoire dAudiologie ExpPrimentale, INSERM unit6 229. Universit6 Bordeaux II, HGpital Pellegrin, 33076 Bordeaux, France
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Lapeyre P,Guilhaume A, Cazals Y.Differences in hair bundles associated with type I and type I1 vestibular hair cells of the guinea pig saccule. Acta Otolaryngol (Stockh) 1992; 112: 635-642. Several studies have reported variations in shape and size of stereociliary bundles and in a limited number of observations have associated them to type I and type I1 hair cells. A systematic study has been undertaken for which a technique was developed in order to identify both cell types and their corresponding hair bundles. Numerous fissures were obtained in saccular epithelia and observed in scanning electron microscopy. Sacculartype I and type I1 hair cells in the guinea pig were found to have distinctive hair bundles. The tallest stereocilia of almost all type I cells were longer than 6 pm, and were shorter in the striola compared to the periphery. In contrast, the tallest stereocilia of almost all type I1 cells were shorter than 6 pm and were not found to vary notably in size from the striola to the periphery. Hair bundles with stereocilia organized in straight or in staggered rows were found for both types of cells across the whole saccular epithelium, with no apparent particular distribution. Possible physiological significanceof differences in hair bundles is discussed. Key words: guinea pig, sacmle. hair cells, stereocilia, scanning electron microscopy.
INTRODUCTION Based on cell-body and synapse morphology, two types of sensory cells have been identified in the vestibular epithelia of higher vertebrates: the amphora-shaped type I hair cells, which are surrounded by an afferent nerve calyx, and the cylinder-shaped type I1 hair cells, which are contacted by neural boutons and are phylogenetically more ancient (1). The vestibular end organs possess hair cells which serve as mechanoreceptors by detecting a stirnulatory force through their hair bundle. These hair tufts are composed of stereocilia and of a kinocilium on one side of the bundle (2). In mammals, they are not only overall shorter in the striolar or central areas and longer in the periphery (3), they also differ greatly in size among the cells of a single region of the epithelium. In addition, different types of hair bundles have been mentioned according to the proportional lengths of the kinocilium and taller stereocilia (4) and to the slope of the stereocilia size gradation ( 5 ) ; the geometrical arrangement of the stereocilia in a hair tuft has also been documented (6). A distinctive functional role of the two kinds of hair cells is still unknown and extensive studies are conducted on mammalian vestibular peripheral innervation patterns (7)and their relation with vestibular afferent discharge properties (S), and also recently on the different membrane conductances of the vestibular type I and type I1 hair cells (9). One significant feature in this perspective could be different ciliary tuft configurations associated with different hair cells. Indeed, it has been suggested that hair bundles with tall stereocilia belong to type I hair cells, while those with shorter stereocilia belong to type I1 cells (10). Fortuitous fractures through rat maculae allowed visualization of type I and type I1 cells side by side that led to the statement that finer stereocilia belonged to type I1 hair cells (5). Since reports in the literature about hair bundle types and associated cell types in the vestibular end organs are based on rather seldom observations (3,we set out to determine, in the guinea pig, whether cell types are associated systematically with particular hair tuft
536 P . Lupeyre et al.
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features, using scanning electron microscopy. In order to identify both the cell types and their corresponding hair bundle, we developed a technique to obtain numerous fissures in saccular epithelia during the critical-point drying stage.
Downloaded by [Universität Osnabrueck] at 07:41 28 January 2016
MATERIAL AND METHODS Bullae from 9 pigmented adult guinea pigs (250-350 g) were used in this study. After dissection in Hank‘s balanced salt solution (Sigma), the otolithic organs were treated by immersion in a fixative containing 2.5% glutaraldehyde buffered with 0.01 M cacodylate for 30 min. The specimens were rinsed i? 0.075 M cacodylate buffer (pH 7.35 and 300 mosM), and the otolithic membrane was removed mechanically with a fine jet of buffer solution or with fine forceps. The organs were then post-fixed with 1 % OsO, for 10 min. After dehydration in graded ethanols, the specimens were dried by the critical-point method (COz;Balzers CPDOIO), mounted on stubs with double-coated sticky tape (Fullam) and covered with a thin layer of gold-palladium in an ion sputter coating unit (Balzers SCD 030). A scanning electron microscope (Philips 505) was used for specimen observation. Special attention was devoted to obtaining sections in the epithelia that would allow observation of both hair cell bodies and their ciliary tuft. Tearing apart the epithelia with fine forceps proved difficult when they were attached to supporting tissue. When they were isolated, pieces of the epithelia were extremely fragile and difficult to fix on stubs. The edges obtained after cutting the epithelia with a rasor blade or a sharp scalpel were curved inwards and therefore did not allow observation of the cell bodies. The technique that proved to be successful was to keep saccules on their underlying bone. Indeed, the critical-point drying stage led to numerous fissures in the epithelia, due to the difference in rigidity and constitution between the sensory tissue and the bone. We therefore used this technique for the rest of the study, using saccules and not utricles as the latter are not attached to the bone. Scanning electron miscroscopy observations were made of cell bodies that could be identified in fissures in the saccular maculae, and of their corresponding hair bundle after adequate orientation for identification. The hair tufts were examined with regard to specially geometrical features and length of the taller stereocilia, measured with a curvometer on micrographs. Furthermore, the spatial distribution of these cells on the corresponding epithelium was recorded. RESULTS Fissures obtained in saccular epithelia during the critical-point drying stage allowed viewing and subsequent identification of hair cell bodies when they were at least 10 pm wide (Fig. 1). Hair cells which had a constricted neck region below the cuticular plate and a spherical base were easily identified as being type I cells, and usually had their cell body covered with debris, probably remnants of the nervous calyx (Fig. 3). Those without a neck region and a more cylindrical shape were identified as type I1 cells, and were often found to have a smooth cell body to which were sometimes attached remnants of neighbouring cells’ membrane (Fig. 4). Among the 162 cells identified in this study, 96 were type I cells and 66 type I1 cells. Our observations indicate that hair tufts from type I cells are clearly taller than those from type I1 cells. The stereocilia of the tallest row of the hair bundle of type I hair cells measured from 5.4 to 14.6 pm, with a mean of 8.9 pm. Those of the type I1 hair cells were much shorter and measured from 2 to 8.9 pm, with a mean of 4.4 pm. In Fig. 2, distributions of hair lengths are given and it appears that almost all type 11 cells’ hair bundle sizes are below 6 pm whereas almost all the hair bundle sizes of type I cells’ are above this value. For type I cells the length of the tallest stereocilia gradually increased from the striola to the
Hair bundles of vestibular type I and 11 cells 637
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Acta Otolarynpl (Slockh) I12
Fig. I . ((I) and (c), micrographs of slceular epithelia showing fissures obtained during the criticalpoint drying stage. (b) and (d), diagrams of locations of type I (circles) and type I1 (sqwa)hair cells showing stereocilia in straight rows ( d i d symbdc), in rows ( o p n s y d o k ) , or non dessifuble (Mf did symbols) hair bundles in tbe sa.ecul.r epitheliaviewed in (u)and (c), rrrpaivdy. (e) and (fi,lLpe I (I) and type11 (1I)hairdls viewed in f m . ScaleLurs represent 1 mm in(@, 0.1 mm in@, c a d &and I Om in (c) and
tn.
Act0 Otolaryngol (Stockh) 112
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Fig.1.((I) and (c), cell body ofa type 11 hair cell viewed in a fissure in the saccular epithelium; (b) and ( d ) , corresponding hair bundle with stercocilia arranged in straight and staggered rows. Scale bars nprcscnt 10 pn in (a)and (c) and 1 w in (b) and (d).
than 6 Bm,whereas the longest stereocilia of almost all type I1 hair cells’ were shorter than so. For type I hair cells only. stereocilialength increased from the striola towards the periphery of the sensory epithelium. Straight or staggered stereocilia rows, previously described in another study as tight and loose (61, were found for both types of hair cells, across all of the saccular epithelium.
Hair brmdles of vestibulm type I cmd II ceh 639
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Fig. 3. ( a ) and (c), cell body of a typc I hair cell viewed in a fissure in the saccular epithelium; (b)and (d), corresponding hair bundle with stereocilia arranged in straight and staggered rows. Scale bars represent 10 pm in (a) and (c) and 1 pm in (b) and ( d ) .
were often tortuous. However, very long Linocilia were found in both types I and I1 hair cells, and appeared generally taller in the former than in the latter. DISCUSSION Data from this study indicate that saccuIar type I and type I1 hair cells in the guinea pig have distinctive hair bundle types: the longest stereocilia of almost all type I hair cells were greater
et d.
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Fig.1.(a)and (c), cell body of a type Il hair cell viewed in a fissure in the saccular epithelium; (b)and (d), corresponding hair bundle with stereocilia arranged in straight and staggered rows. Scale bars represent 10 pm in (a)and (c) and 1 pm in (b) and (d).
than 6 pm, whereas the longest stereocilia of almost all type I1 hair cells’ were shorter than so. For type I hair cells only, stereocilia length increased from the striola towards the periphery of the sensory epithelium. Straight or staggered stereocilia rows, previously described in another study as tight and loose (6), were found for both types of hair cells, across all of the saccular epithelium.
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While with transmission electron microscopy it is not easy to observe the whole length of stereocilia bundles in the plane of a transverse section, and it is impossible to tell whether their stereocilia are organized in straight or in staggered rows, in scanning electron microscopy it is not possible to identify the type of sensory cells, except in exceptional fortuitous fractures (5, 14). Our technique succeeded in obtaining numerous and large fractures in the same organ. To identify the type of hair cells, our main criterion was the cell shape, and sometimes the smoothness of the cell body. We found that some type I1 cells were not very cylindrical and had either a short or tall cell body, as described elsewhere for isolated cells from the cristae ampullares (9). The precise percentage of type I and type I1 cells in the mammalian otolithic organs has not been studied thoroughly as in the cristae ampullares (1 5). We found that both types of cells were present throughout the epithelium, in accordance with previous observations (3). Our results indicated that only for type I cells hair bundles stereocilia are shorter in the striola than in the periphery and thus restrict previous global statements (3, 4). The longest stereocilia from our data measured nearly 15 Fm, in accordance with previous observations (3) and discordance with others (10). We did not find a particular distribution of hair bundles with stereocilia in straight or in staggered rows between the two types of hair cells. Moreover, we did not find the same gradient as in another study (6) for these two types of hair bundles across the epithelium. Speculations have previously been made about the way the different stereocilia types could be linked or not to the otoconial membrane in the otolithic organs, in parallel with what is found in the cochlea (16), so as to find a different role of the type I and type I1 hair cells and to possibly relate to the different dynamic properties of vestibular afferent fibers (10, 17). However, it is well established in mammals, as well as in low vertebrates where only type I1 cells exist but present different types of stereociliary bundles, that the different firing properties of vestibular afferents are more closely correlated to their epithelial location than to the type and number of hair cells they contact (18, 19). These differences in afferent properties could reflect the morphology and the coupling of the cells’ hair bundles to the otoconial membrane. Indeed, longer hair bundles have been shown to be less sensitive to mechanical stimulation in the chick (20). As we found that the type 1 hair cells hair bundles in the periphery are long, we can speculate, in line with another author (lo), that the tip of their kinocilium and taller stereocilia could be linked to the otoconial membrane and that these cells therefore could respond preferably to displacement, while the hair bundles from the type I1 hair cells could be freestanding as they are shorter and would therefore respond to velocity. In the striola where the stereocilia are relatively short compared t o the periphery, and where wide holes are present in the otoconial membrane (3) the hair bundles could be free standing, and the cells would therefore be more velocity receptive in this region. However, the type of linkage of the stereocilia to the otolithic membrane is still an unresolved problem as the existence of a space between the sensory epithelium and the accessory membrane is still a matter of dispute (3). ACKNOWLEDGEMENTS The authors thank Dr Jonathan F. Ashmore for carefully reading and improving the manuscript and JCrome Dupont for his assistance with the figures. This work was supported by a grant to the first author from the Ministere de la Recherche et de la Technologie.
REFERENCES 1. WersZll J. Studies on the structure and innervation of the sensory epithelium of the cristae ampullares
in the guinea pig. A light and electron microscopicinvestigation. Acta Otolaryngoi (Stockh) 1956 Suppl 126.
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2. Hudspeth AJ, Corey DP. Sensitivity, polarity and conductance change in the response of vertebrate hair cells to controlled mechanical stimuli. Proc Natl Acad Sci USA 1977; 74: 2407-1 1. 3. Lindeman HH. Anatomy of the otolithic organs. Adv Otorhinolaryngol 1973; 2 0 405-33. 4. Lim DJ, Morphological and physiological correlates in cochlear and vestibular sensory epithelia. Scand Electron Microsc 1976; 5: 269-76. 5. Ross MD, Donovan K, Rogers C. Peripheral sensory processing in mammalian gravity receptors: observations of ciliary tuft configurations. In: Graham MD, Kemink JL, eds. The vestibular system: neurophysiologic and clinical research. New York Raven Press, 1987: 119-24. 6. Bagger-Sjoback D, Takumida M. Geometrical array of the vestibular sensory hair bundle. Acta Otolaryngol (Stockh) 1988; 106 393-403. 7. Ross MD, Rogers CM, Donovan KM. Innervation patterns in rat saccular macula. Acta Otolaryngol (StWkh) 1986; 1 0 2 75-86. 8. Goldberg JM, Fernindez C. Physiology of peripheral neurons innervating semicircular canals of the squirrel monkey. Ill. Variations among units in their discharge properties. J Neurophysiol 1971; 34: 676-84. 9. Rennie KJ, Ashmore JF. Ionic currents in isolated vestibular hair cells from the guinea-pig crista ampullaris. Hear Res 1991; 51: 279-92. 10. Lim DJ. Ultra anatomy of sensory end-organs in the labyrinth and their functional implications. In: Shambaugh GE Jr, Shea JJ, eds. Proceedings of the Shambaugh 5th international workshop on middle ear microsurgery and fluctuant hearing loss. Huntville: Strode Publ, 1977: 16-27. 11. Lim DJ, Anniko M. Developmental morphology of the mouse inner ear. A S.E.M. observation. Acta Otolaryngol (Stockh) 1985; Suppl422. 12. Femandez C, Baird RA, Goldberg JM. The vestibular nerve of the chinchilla. I. Peripheral innervation patterns in the horizontal and superior semicircular canals. J Neurophysiol 1988; 6 0 167-81. 13. Fernandez C, Goldberg JM, Baird RA. The vestibular nerve of the chinchilla. 111. Peripheral innervation patterns in the utricular macula. J Neurophysiol 1990; 63: 767-80. 14. Hunter-Duvar 1M. An electron microscopic study of the vestibular sensory epithelium. Acta Otolaryngo1 (Stockh) 1983; 95: 494-507. 15. Lindeman HH, Reith A, Winther F0. The distribution of type I and type I1 cells in the cristae ampullares of the guinea pig. A morphometric investigation. Acta Otolaryngol (Stockh) 1981; 92: 315-21. 16. Dallos P, Billone MC, Durrant JD. Cochlear inner and outer hair cells: functional differences. Science 1972; 177: 356-8. 17. Goldberg JM, Femindez C. Vestibular mechanisms. Ann Rev Physiol 1975; 37: 129-62. 18. Baird RA,Lewis ER. Correspondencesbetween afferent innervation patterns and response dynamics in the bullfrog utricle and lagena. Brain Res 1986; 3 6 9 48-64. 19. Goldberg JM, Desmadryl G, Baird RA, Fernindez C. The vestibular nerve of the chinchilla. V. Relation between afferent discharge properties and peripheral innervation patterns in the utricular macula. J Neurophysiol 1990; 63: 791-804. 20. Ohmori H. Gating properties of the mechano-electricaltransducer channel in the dissociated vestibular hair cell of the chick. J Physiol (Lond) 1987; 387: 589-609.
Manuscript received July 22. 1991; accepted Augusi 22, 1991 Address for correspondence: Yves Cazals, Laboratoire d'Audiologie Exp&imentale, INSERM unitt 229, Universitk de Bordeaux 11, Hbpital Pellegrin, F-33076 Bordeaux, France
ISSN: 0001-6489 (Print) 1651-2251 (Online) Journal homepage: http://www.tandfonline.com/loi/ioto20
Differences in Hair Bundles Associated with Type I and Type II Vestibular Hair Cells of the Guinea Pig Saccule Pascale Lapeyre, Anne Guilhaume & Yves Cazals To cite this article: Pascale Lapeyre, Anne Guilhaume & Yves Cazals (1992) Differences in Hair Bundles Associated with Type I and Type II Vestibular Hair Cells of the Guinea Pig Saccule, Acta Oto-Laryngologica, 112:4, 635-642 To link to this article: http://dx.doi.org/10.3109/00016489209137453
Published online: 08 Jul 2009.
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Article views: 5
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Full Terms & Conditions of access and use can be found at http://www.tandfonline.com/action/journalInformation?journalCode=ioto20 Download by: [Universität Osnabrueck]
Date: 28 January 2016, At: 07:41
Acta Otolaryngol (Stockh) 1992; 112: 635-642
Differences in Hair Bundles Associated With Type I and Type I1 Vestibular Hair Cells of the Guinea Pig Saccule PASCALE LAPEYRE, ANNE GUILHAUME and YVES CAZALS From the Laboratoire dAudiologie ExpPrimentale, INSERM unit6 229. Universit6 Bordeaux II, HGpital Pellegrin, 33076 Bordeaux, France
Downloaded by [Universität Osnabrueck] at 07:41 28 January 2016
Lapeyre P,Guilhaume A, Cazals Y.Differences in hair bundles associated with type I and type I1 vestibular hair cells of the guinea pig saccule. Acta Otolaryngol (Stockh) 1992; 112: 635-642. Several studies have reported variations in shape and size of stereociliary bundles and in a limited number of observations have associated them to type I and type I1 hair cells. A systematic study has been undertaken for which a technique was developed in order to identify both cell types and their corresponding hair bundles. Numerous fissures were obtained in saccular epithelia and observed in scanning electron microscopy. Sacculartype I and type I1 hair cells in the guinea pig were found to have distinctive hair bundles. The tallest stereocilia of almost all type I cells were longer than 6 pm, and were shorter in the striola compared to the periphery. In contrast, the tallest stereocilia of almost all type I1 cells were shorter than 6 pm and were not found to vary notably in size from the striola to the periphery. Hair bundles with stereocilia organized in straight or in staggered rows were found for both types of cells across the whole saccular epithelium, with no apparent particular distribution. Possible physiological significanceof differences in hair bundles is discussed. Key words: guinea pig, sacmle. hair cells, stereocilia, scanning electron microscopy.
INTRODUCTION Based on cell-body and synapse morphology, two types of sensory cells have been identified in the vestibular epithelia of higher vertebrates: the amphora-shaped type I hair cells, which are surrounded by an afferent nerve calyx, and the cylinder-shaped type I1 hair cells, which are contacted by neural boutons and are phylogenetically more ancient (1). The vestibular end organs possess hair cells which serve as mechanoreceptors by detecting a stirnulatory force through their hair bundle. These hair tufts are composed of stereocilia and of a kinocilium on one side of the bundle (2). In mammals, they are not only overall shorter in the striolar or central areas and longer in the periphery (3), they also differ greatly in size among the cells of a single region of the epithelium. In addition, different types of hair bundles have been mentioned according to the proportional lengths of the kinocilium and taller stereocilia (4) and to the slope of the stereocilia size gradation ( 5 ) ; the geometrical arrangement of the stereocilia in a hair tuft has also been documented (6). A distinctive functional role of the two kinds of hair cells is still unknown and extensive studies are conducted on mammalian vestibular peripheral innervation patterns (7)and their relation with vestibular afferent discharge properties (S), and also recently on the different membrane conductances of the vestibular type I and type I1 hair cells (9). One significant feature in this perspective could be different ciliary tuft configurations associated with different hair cells. Indeed, it has been suggested that hair bundles with tall stereocilia belong to type I hair cells, while those with shorter stereocilia belong to type I1 cells (10). Fortuitous fractures through rat maculae allowed visualization of type I and type I1 cells side by side that led to the statement that finer stereocilia belonged to type I1 hair cells (5). Since reports in the literature about hair bundle types and associated cell types in the vestibular end organs are based on rather seldom observations (3,we set out to determine, in the guinea pig, whether cell types are associated systematically with particular hair tuft
536 P . Lupeyre et al.
Acta Otolaryngol (Stockh) 112
features, using scanning electron microscopy. In order to identify both the cell types and their corresponding hair bundle, we developed a technique to obtain numerous fissures in saccular epithelia during the critical-point drying stage.
Downloaded by [Universität Osnabrueck] at 07:41 28 January 2016
MATERIAL AND METHODS Bullae from 9 pigmented adult guinea pigs (250-350 g) were used in this study. After dissection in Hank‘s balanced salt solution (Sigma), the otolithic organs were treated by immersion in a fixative containing 2.5% glutaraldehyde buffered with 0.01 M cacodylate for 30 min. The specimens were rinsed i? 0.075 M cacodylate buffer (pH 7.35 and 300 mosM), and the otolithic membrane was removed mechanically with a fine jet of buffer solution or with fine forceps. The organs were then post-fixed with 1 % OsO, for 10 min. After dehydration in graded ethanols, the specimens were dried by the critical-point method (COz;Balzers CPDOIO), mounted on stubs with double-coated sticky tape (Fullam) and covered with a thin layer of gold-palladium in an ion sputter coating unit (Balzers SCD 030). A scanning electron microscope (Philips 505) was used for specimen observation. Special attention was devoted to obtaining sections in the epithelia that would allow observation of both hair cell bodies and their ciliary tuft. Tearing apart the epithelia with fine forceps proved difficult when they were attached to supporting tissue. When they were isolated, pieces of the epithelia were extremely fragile and difficult to fix on stubs. The edges obtained after cutting the epithelia with a rasor blade or a sharp scalpel were curved inwards and therefore did not allow observation of the cell bodies. The technique that proved to be successful was to keep saccules on their underlying bone. Indeed, the critical-point drying stage led to numerous fissures in the epithelia, due to the difference in rigidity and constitution between the sensory tissue and the bone. We therefore used this technique for the rest of the study, using saccules and not utricles as the latter are not attached to the bone. Scanning electron miscroscopy observations were made of cell bodies that could be identified in fissures in the saccular maculae, and of their corresponding hair bundle after adequate orientation for identification. The hair tufts were examined with regard to specially geometrical features and length of the taller stereocilia, measured with a curvometer on micrographs. Furthermore, the spatial distribution of these cells on the corresponding epithelium was recorded. RESULTS Fissures obtained in saccular epithelia during the critical-point drying stage allowed viewing and subsequent identification of hair cell bodies when they were at least 10 pm wide (Fig. 1). Hair cells which had a constricted neck region below the cuticular plate and a spherical base were easily identified as being type I cells, and usually had their cell body covered with debris, probably remnants of the nervous calyx (Fig. 3). Those without a neck region and a more cylindrical shape were identified as type I1 cells, and were often found to have a smooth cell body to which were sometimes attached remnants of neighbouring cells’ membrane (Fig. 4). Among the 162 cells identified in this study, 96 were type I cells and 66 type I1 cells. Our observations indicate that hair tufts from type I cells are clearly taller than those from type I1 cells. The stereocilia of the tallest row of the hair bundle of type I hair cells measured from 5.4 to 14.6 pm, with a mean of 8.9 pm. Those of the type I1 hair cells were much shorter and measured from 2 to 8.9 pm, with a mean of 4.4 pm. In Fig. 2, distributions of hair lengths are given and it appears that almost all type 11 cells’ hair bundle sizes are below 6 pm whereas almost all the hair bundle sizes of type I cells’ are above this value. For type I cells the length of the tallest stereocilia gradually increased from the striola to the
Hair bundles of vestibular type I and 11 cells 637
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Acta Otolarynpl (Slockh) I12
Fig. I . ((I) and (c), micrographs of slceular epithelia showing fissures obtained during the criticalpoint drying stage. (b) and (d), diagrams of locations of type I (circles) and type I1 (sqwa)hair cells showing stereocilia in straight rows ( d i d symbdc), in rows ( o p n s y d o k ) , or non dessifuble (Mf did symbols) hair bundles in tbe sa.ecul.r epitheliaviewed in (u)and (c), rrrpaivdy. (e) and (fi,lLpe I (I) and type11 (1I)hairdls viewed in f m . ScaleLurs represent 1 mm in(@, 0.1 mm in@, c a d &and I Om in (c) and
tn.
Act0 Otolaryngol (Stockh) 112
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Fig.1.((I) and (c), cell body ofa type 11 hair cell viewed in a fissure in the saccular epithelium; (b) and ( d ) , corresponding hair bundle with stercocilia arranged in straight and staggered rows. Scale bars nprcscnt 10 pn in (a)and (c) and 1 w in (b) and (d).
than 6 Bm,whereas the longest stereocilia of almost all type I1 hair cells’ were shorter than so. For type I hair cells only. stereocilialength increased from the striola towards the periphery of the sensory epithelium. Straight or staggered stereocilia rows, previously described in another study as tight and loose (61, were found for both types of hair cells, across all of the saccular epithelium.
Hair brmdles of vestibulm type I cmd II ceh 639
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Fig. 3. ( a ) and (c), cell body of a typc I hair cell viewed in a fissure in the saccular epithelium; (b)and (d), corresponding hair bundle with stereocilia arranged in straight and staggered rows. Scale bars represent 10 pm in (a) and (c) and 1 pm in (b) and ( d ) .
were often tortuous. However, very long Linocilia were found in both types I and I1 hair cells, and appeared generally taller in the former than in the latter. DISCUSSION Data from this study indicate that saccuIar type I and type I1 hair cells in the guinea pig have distinctive hair bundle types: the longest stereocilia of almost all type I hair cells were greater
et d.
Act8 OcoLrYllgol ~Stodrh)I12
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640 P. Lapyre
Fig.1.(a)and (c), cell body of a type Il hair cell viewed in a fissure in the saccular epithelium; (b)and (d), corresponding hair bundle with stereocilia arranged in straight and staggered rows. Scale bars represent 10 pm in (a)and (c) and 1 pm in (b) and (d).
than 6 pm, whereas the longest stereocilia of almost all type I1 hair cells’ were shorter than so. For type I hair cells only, stereocilia length increased from the striola towards the periphery of the sensory epithelium. Straight or staggered stereocilia rows, previously described in another study as tight and loose (6), were found for both types of hair cells, across all of the saccular epithelium.
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Acta Otolaryngol (Stockh) 112
Hair bundles of vestibular type I and II cells 641
While with transmission electron microscopy it is not easy to observe the whole length of stereocilia bundles in the plane of a transverse section, and it is impossible to tell whether their stereocilia are organized in straight or in staggered rows, in scanning electron microscopy it is not possible to identify the type of sensory cells, except in exceptional fortuitous fractures (5, 14). Our technique succeeded in obtaining numerous and large fractures in the same organ. To identify the type of hair cells, our main criterion was the cell shape, and sometimes the smoothness of the cell body. We found that some type I1 cells were not very cylindrical and had either a short or tall cell body, as described elsewhere for isolated cells from the cristae ampullares (9). The precise percentage of type I and type I1 cells in the mammalian otolithic organs has not been studied thoroughly as in the cristae ampullares (1 5). We found that both types of cells were present throughout the epithelium, in accordance with previous observations (3). Our results indicated that only for type I cells hair bundles stereocilia are shorter in the striola than in the periphery and thus restrict previous global statements (3, 4). The longest stereocilia from our data measured nearly 15 Fm, in accordance with previous observations (3) and discordance with others (10). We did not find a particular distribution of hair bundles with stereocilia in straight or in staggered rows between the two types of hair cells. Moreover, we did not find the same gradient as in another study (6) for these two types of hair bundles across the epithelium. Speculations have previously been made about the way the different stereocilia types could be linked or not to the otoconial membrane in the otolithic organs, in parallel with what is found in the cochlea (16), so as to find a different role of the type I and type I1 hair cells and to possibly relate to the different dynamic properties of vestibular afferent fibers (10, 17). However, it is well established in mammals, as well as in low vertebrates where only type I1 cells exist but present different types of stereociliary bundles, that the different firing properties of vestibular afferents are more closely correlated to their epithelial location than to the type and number of hair cells they contact (18, 19). These differences in afferent properties could reflect the morphology and the coupling of the cells’ hair bundles to the otoconial membrane. Indeed, longer hair bundles have been shown to be less sensitive to mechanical stimulation in the chick (20). As we found that the type 1 hair cells hair bundles in the periphery are long, we can speculate, in line with another author (lo), that the tip of their kinocilium and taller stereocilia could be linked to the otoconial membrane and that these cells therefore could respond preferably to displacement, while the hair bundles from the type I1 hair cells could be freestanding as they are shorter and would therefore respond to velocity. In the striola where the stereocilia are relatively short compared t o the periphery, and where wide holes are present in the otoconial membrane (3) the hair bundles could be free standing, and the cells would therefore be more velocity receptive in this region. However, the type of linkage of the stereocilia to the otolithic membrane is still an unresolved problem as the existence of a space between the sensory epithelium and the accessory membrane is still a matter of dispute (3). ACKNOWLEDGEMENTS The authors thank Dr Jonathan F. Ashmore for carefully reading and improving the manuscript and JCrome Dupont for his assistance with the figures. This work was supported by a grant to the first author from the Ministere de la Recherche et de la Technologie.
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Manuscript received July 22. 1991; accepted Augusi 22, 1991 Address for correspondence: Yves Cazals, Laboratoire d'Audiologie Exp&imentale, INSERM unitt 229, Universitk de Bordeaux 11, Hbpital Pellegrin, F-33076 Bordeaux, France
