Cell Tiss. Res. 190, 115-121 (1978)
Cell and Tissue Research 9 by Springer-Verlag 1978
Gap Junctions in Human Sebaceous Glands Neil Kitson, E.W. Van Lennep and J.A. Y o u n g * Departments of Physiology, and of Histology and Embryology, University of Sydney, Sydney,Australia Summary. A n examination o f h u m a n sebaceous glands by transmission electron m i c r o s c o p y has revealed the presence o f gap junctions. The junctions are f o u n d in a b u n d a n c e between differentiating cells, and annular forms are also seen. The possible significance o f this new finding is briefly discussed.
Key words: Sebaceous gland - H u m a n - G a p junctions.
Introduction Several authors have c o m m e n t e d on intercellular connections in h u m a n sebaceous glands (Charles, 1960; Hibbs, 1962; Ellis and Henrikson, 1963; Ellis, 1967; Fujita et al., 1972). While desmosomes have been reported in all these investigations, only two authors have published micrographs o f m e m b r a n e s in close apposition. Ellis (1967) c o m m e n t e d on the existence o f an "... occasional tight junction or nexus" and Bell (1974) noted "double walled structures" which she speculated were " p r o b a b l y specialized junctions". In our study o f h u m a n sebaceous glands, we have f o u n d m e m b r a n e s in close apposition having the characteristic appearance and dimensions o f gap junctions. These structures have been f o u n d in every gland so far examined.
Materials and Methods Tissue
Normal human skin was retrieved from ENT surgery in accordance with a protocol approved by the Medical Ethics Committees of the University of Sydney, and the Royal Prince Alfred Hospital, Sydney. Send offprint requests to: Dr. N. Kitson, Dept. of Physiology, University of Sydney, NSW 2006, Australia * We thank the ENT consultant staff of Royal Prince Alfred Hospital, Sydney, for making tissue available. The co-operation of the director, Dr. D.J.H. Cockayne, and staff of the University of Sydney Electron Microscope Unit is gratefully acknowledged. Mr. E. Foster provided excellent assistance with the photography. - The project was supported by the Consolidated Medical Research Funds, University of Sydney, and the National Health and Medical Research Council of Australia. One of us (N.K.) was the recipient of the Phyllis Anderson Medical Research Fellowship awarded by the Faculty of Medicine, University of Sydney
0302 766X/78/0190/0115/$01.40
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Samples were obtained in all cases from patients requiring post-auricular incisions.A strip of skin 1 mm wide was removed from the wound edge. This procedure had no influenceon the size or location of the initial incision.
Thin Sections
Excised skin was immediatelyimmersedin 2.5 % glutaraldehyde buffered with 0.1 M sodium cacodylate to pH 7.2. After 24 h primary fixation, samples were washed twice in buffer, post-fixed in cacodylatebuffered OsO~ for 2 h, block-stained for 45 min with 0.5 % uranyl acetate, rapidly dehydrated in an acetone series to a 50/50 mixture of Spurr's resin and acetone, and left overnight. Infiltration was accomplished over 48 h in two changes of 100 % resin, and the blocks were then embedded. Grey-to-silversectionswere cut on a Reichert OMU-3 ultramicrotome and mounted on naked, 400 mesh, copper grids. After staining with uranyl acetate and Reynolds' lead (Reynolds,1963), sectionswere viewed in a Philips 201 electron microscope operating at 60 KV with a 50 mcm objective aperture.
Lanthanum
Some tissue blocks were processed with lanthanum hydroxide present in all solutions up to the dehydration sequence after the method of Revel and Karnovsky (1967). Otherwise, the tissue was handled as for thin sections.
Results G a p j u n c t i o n s could be seen at lower powers (Figs. 1-3) as dense bands. A t higher powers (Figs. 4, 6), the characteristic "seven layered" structure ( B r i g h t m a n a n d Reese, 1969) became apparent, a n d the gap was discernible as were periodic crossstriations. The j u n c t i o n widths m e a s u r e d between 1 6 . 5 - 1 8 . 0 n m in r a n d o m m e a s u r e m e n t s with a n optical micrometer. The gap widths were a b o u t 2 - 4 nm, a n d l a n t h a n u m infiltrated j u n c t i o n s (Fig. 5) showed a dense central b a n d 4 - 6 n m wide. T h e l a n t h a n u m could n o t be distinguished from the electron-dense outer leaflets of the a p p o s i n g m e m b r a n e s . These results are in general agreement with those of other a u t h o r s (Staehelin, 1974). The hexagonal p a t t e r n usually seen in oblique sections of l a n t h a n u m - t r e a t e d j u n c t i o n s (Revel a n d K a r n o v s k y , 1967) has n o t been observed in our preparations. A n n u l a r j u n c t i o n s were occasionally seen (Fig. 6). The area enclosed by the j u n c t i o n always a p p e a r e d less electron-dense t h a n the s u r r o u n d i n g cytoplasm, b u t n o recognizable organelles were seen within the j u n c t i o n s . We have n o t attempted to determine whether or n o t a c o n n e c t i o n exists between these structures a n d the cytoplasmic m e m b r a n e , b u t so far a n n u l a r j u n c t i o n s have n o t been seen in l a n t h a n u m - t r e a t e d tissue. A l t h o u g h a q u a n t i t a t i v e analysis c a n n o t be offered, a n impression has been gained that gap j u n c t i o n s are f o u n d almost entirely between differentiating sebaceous cells, a n d n o t between undifferentiated or fully differentiated cells.
G a p Junctions in H u m a n Sebaceous Glands
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Fig. 1. Sebaceous cells from a 13 year old male ( x 15,000). Arrows show two gap junctions in apposing membranes of differentiating cells. L lipid Fig. 2. Sebaceous cells from a 13 year old male ( x 30,000). A n extensive junction is seen between the arrows. L lipid Fig.3. Sebaceous cells from a 13 year old male ( x 25,000). Arrow marks small junction
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Fig.4. Sebaceous cells from a 13 year old male ( x 236,000). The gap is visible as are periodic cross striations Fig. 5. Sebaceous cells from a 44 year old male ( x 261,000). L a n t h a n u m has infiltrated the gap and obscured the electron dense outer leaflets. Periodic electron lucent areas are faintly visible Fig. 6. Sebaceous cells from a 13 year old female ( x 79,000). A n annular junction is shown with enclosed area appearing less electron dense than surrounding cytoplasm. Inset ( • 299,000) shows gap in same junction
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This is the first report of gap junctions in sebaceous glands although the structures previously reported by Ellis (1967) and Bell (1974) were undoubtedly also gap junctions. Such junctions have been observed in wool follicles (Orwin et al., 1973), rat oral mucosa (Shimono and Clementi, 1976) and human epidermis (Wolff and Schreiner, 1968; Hashimoto, 1971; Vreeswijk et al., 1977). Cavoto and Flaxman (1972) found gap junctions between epidermal cells in culture, and also established the existence of low-resistance electrical connections between such cells. Gap junction subunits are thought to be intercellular channels (Revel and Karnovsky, 1967; Brightman and Reese, 1969; Peracchia, 1973; Caspar et al., 1977; Makowski et al., 1977) which allow electrical coupling (Loewenstein and Kanno, 1964; Rose, 1971; Ito et al., 1974) and metabolic co-operation (Gilula et al., 1972; Azarnia et al., 1974; Pitts and Simms, 1977). Electrical coupling has been observed in several exocrine glands that respond to autonomic stimulation (Ginsborg et al., 1974; Petersen and Ueda, 1976), but the role of membrane conductance changes in secretion is not clear. Lawrence et al. (1977) have evidence to suggest that intra cellular messengers are transmitted through gap junctions; membrane conductance changes in coupled cells may conceivably be secondary to such phenomena. In the case of sebaceous glands, secretion involves cell lysis (Palay, 1958) after several days' differentiation (Downing et al., 1975). In addition, the glands are said not to be innervated (Montagna, 1963), and thus close autonomic control of secretion by regulation of membrane conductance is unlikely; exchange of chemical information through the junctions is more probable. However, Dugan (1974) and F6rster (1975) have reported autonomic nerves innervating sebaceous gland basal cells respectively, in rat and mouse, and the question must remain open. "Annular" gap junctions have been observed in other tissues (Garant et al., 1972; Merk et al., 1973; Orwin et al., 1973; Bjersing and Cajander, 1974; Albertini et al., 1975), in tissue culture (Perissel et al., 1976), in tumour (Letourneau et al., 1975), and in epidermis (Vreeswij k et al., 1977). By serial sectioning, Vreeswijk et al. showed that the junction limited vesicles so studied were not connected to cytoplasmic membrane. In other material however, Merk et al. (1973), Letourneau et al. (1975), and Perissel et al. (1976) showed that lanthanum infiltrated some annular junctions, implying continuation with the intercellular space. Van Lennep and Madden (1965) observed similar membrane-enclosed vesicles in human corpus luteum, and although they described the membranes as"quintuple layered" as found in the zonula occludens, the structures are more likely gap junctions of the annular type. These authors interpreted their findings as a "pinching off' of cytoplasmic membrane, followed by destruction of the enclosed material. Albertini et al. (1975) speculated that such "interiorization" of gap junctions may be a method of removing the restricting influence of these connections on cell movements. Thus, a cell might repeatedly make and break connections with adjacent cells while moving through an epithelium. While van Lennep and Madden, and Albertini et al. were referring to ovarian granulosa cells, the same hypothesis could easily be extended to epidermal and sebaceous cells. Interestingly, the frequency of annular junctions found in estrogen-dependent renal adenocarcinoma appears to decrease when estrogen is withdrawn (Letourneau et al., 1975).
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The finding of gap junctions between sebaceous cells is of interest, although perhaps not unexpected given the prevalence of these connections in epithelia generally. The concept, however, that intercellular communication may play a role in the regulation of this form of holocrine secretion is attractive. It remains to be established that the characteristic hexagonal subunit pattern exists between the membranes of such junctions, and that low resistance pathways are present between cells so connected. References Albertini, D.F., Fawcett, D.W., Olds, P.J.: Morphological variations in gap junctions of ovarian granulosa cells. Tissue and Cell 7, 389-405 (1975) Azarnia, R., Larsen, W.J., Loewenstein, W.R.: The membrane junctions in communicating and noncommunicating cells, their hybrids, and segregants. Proc. nat. Acad. Sci. (Wash.) 71, 880-884 (1974) Bell, M.: A comparative study of the ultrastructure of the sebaceous glands of man and other primates. J. invest. Dermatol. 62, 132-143 (1974) Bjersing, L., Cajander, S.: Ovulation and the mechanism of follicle rupture. IV. Ultrastructure of membrana granulosa of rabbit Graafian follicles prior to induced ovulation. Cell Tiss. Res. 153,1-14 (1974) Brightman, M.W., Reese, T.S.: Junctions between intimately apposed cell membranes in the vertebrate brain. J. Cell Biol. 40, 648~77 (1969) Caspar, D.L.D., Goodenough, D.A., Makowski, L., Phillips, W.C.: Gap junction structures. I. Correlated electron microscopy and X-ray diffraction. J. Cell Biol. 74, 605-628 (1977) Cavoto, F.V., Flaxman, B.A.: Communication between normal human epidermal cells in vitro. J. invest. Dermatol. 59, 370-374 (1972) Charles, A.: Electron microscopic observations of the human sebaceous gland. J. invest. Dermatol. 35, 31-36 (1960) Downing, D.T., Strauss, J.S., Ramasastry, P., Abel, M., Lees, C.W., Pochi, P.E.: Measurement of the time between synthesis and surface excretion of sebaceous lipids in sheep and man. J. invest. Dermatol. 64, 215-219 (1975) Dugan, K.H.: Ultrastructural observations of possible nerve endings in rat sebaceous glands. Cell Tiss. Res. 150, 333-348 (1974) Ellis, R.A.: Eccrine, sebaceous, and apocrine glands. In: Ultrastructure of Normal and Abnormal Skin. 1st ed. (A.S. Zelickson, ed.), pp. 132-162. London: Henry Kimpton 1967 Ellis, R.A., Henrikson, R.C.: The ultrastructure of the sebaceous glands of man. In: Advances in Biology of Skin, Vol. IV (W. Montagna, R.A. Ellis and A.F. Silver, eds.), pp. 94-109. Oxford: Pergamon Press 1963 F6rster, F2., Heine, H., Schaeg, G.: Histophysiology of the vegetative peripheral nervous system of skin. Arch. Dermatol. Res. 254, 295-302 (1975) Fujita, H., Asagami, C., Murata, S., Murozumi, S.: Ultrastructural study of embryonic sebaceous cells, especially of their sebum droplet formation. Acta derm.-venerol. (Stockh.) 52, 99-115 (1972) Garant, P.R.: The demonstration of complex gap junctions between the cells of the enamel organ with lanthanum nitrate. J. Ultrastruct. Res. 40, 333-348 (1972) Gilula, N.B., Reeves, O.R., Steinbach, A.: Metabolic coupling, ionic coupling, and cell contacts. Nature (Lond.) 235, 262-265 (1972) Ginsborg, B.L., House, C.R., Silinsky, E.M.: Conductance changes associated with the secretory potential in the cockroach salivary gland. J. Physiol. (Lond.) 236, 723-731 (1974) Hashimoto, K.: Intercellular spaces of the human epidermis as demonstrated with lanthanum. J. invest. Dermatol. 57, 17-31 (1971) Hibbs, R.G. : Electron microscopy of human axillary sebaceous glands. J. invest. Dermatol. 38, 329-336 (1962) Ito, S., Sato, E., Loewenstein, W.R.: Studies on the formation of a permeable cell membrane junction. II. Evolving junctional conductance and junctional insulation. J. Membrane Biol. 19, 339-355 (1974)
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Lawrence, T.S., Beers, W.H., Gilula, N.B.: Hormonal stimulation and cell communication in cocultures. J. Cell Biol. 75, 63a (1977) Letourneau, R.J., Li, J.J., Rosen, S., Villee, C.A.: Junctional specialization in estrogen-induced renal adenocarcinomas of the golden hamster. Cancer Res. 35, 6-10 (1975) Loewenstein, W.R., Kanno, Y.: Studies on an epithelial cell junction. I. Modifications of surface membrane permeability. J. Cell Biol. 22, 565-586 (1964) Makowksi, L., Caspar, D.L.D., Phillips, W.C., Goodenough, D.A.: Gap junction structures. II. Analysis of the X-ray diffraction data. J. Cell Biol. 74, 629~545 (1977) Merk, F.B., Albright, J.T., Botticelli, C.R. : The fine structure of granulosa cell nexuses in rat ovarian follicles. Anat. Rec. 175, 107-125 (1973) Montagna, W.: The sebaceous glands in man. In:Advances in Biology of Skin. Vol. IV. The Sebaceous Glands, (W. Montagna, R.A. Ellis, and A.F. Silver, eds.), pp. 19-35. Oxford: Pergamon Press 1963 Orwin, D.F.G., Thomson, R.W., Flower, N.E.: Plasma membrane differentiations of keratinizing cell wool follicles. I. Gap junctions. J. Ultrastruct. Res. 45, 1-14 (1973) Palay, S.L.: The morphology of secretion. In: Frontiers in Cytology. (S.L. Palay, ed.), pp. 305-342. New Haven: Yale University Press 1958 Peracchia, C.: Low resistance junctions in crayfish. I. Two arrays of globules in junctional membranes. J. Cell Biol. 57, 54-76 (1973) Perissel, B., Charbonnt, F., Chessebeuf, M., Malet, P.: Differentiation of the plasma membrane of hepatic cells in monolayer cultures. Cell Tiss. Res. 171, 157-173 (1976) Petersen, O.H., Ueda, N.: Pancreatic acinar cells: the role of calcium in stimulus-secretion coupling. J. Physiol. (Lond.) 254, 583-606 (1976) Pitts, J.D., Simms, J.W.: Permeability of junctions between animal cells. Exp. Cell Res. 104, 153-163 (1977) Revel, J.P., Karnovsky, M.I. : Hexagonal array of subunits in intercellular junctions of the mouse heart and liver. J. Cell Biol. 33, C7-C17 (1967) Reynolds, E.S.: The use of lead citrate at high pH as an electron-opaque stain in electron microscopy. J. Cell Biol. 17, 208-213 (1963) Rose, B.: Intercellular communication and some structural aspects of membrane junctions in a simple cell system. J. Membrane Biol. 5, 1-19 (1971) Shimono, M., Clementi, F.: Intercellular connections of oral epithelium. I. Studies with freeze-fracture and tracing methods of normal rat keratinized oral epithelium. J. Ultrastruct. Res. 56, 121-136 (1976) Staehelin, A.: Structure and function of intercellular junctions. Int. Rev. Cytol. 39, 191-283 (1974) Van Lennep, E.W., Madden, L.M.: Electron microscope observations on the involution of the human corpus luteum of menstruation. Z. Zellforsch. 66, 365-380 (1965) Vreeswijk, J., Leene, W., Kalsbeek, G.L.: Early host cell - Molluscum contagiosum virus interactions. II. Viral interactions with the basal epidermal cells. J. invest. Dermatol. 69, 249-256 (1977) Wolff, K., Schreiner, E.: An electron microscopic study on the extraneous coat of keratinocytes and the intercellular space of the epidermis. J. invest. Dermatol. 51, 418430 (1968)
Accepted February 9, 1978
Cell and Tissue Research 9 by Springer-Verlag 1978
Gap Junctions in Human Sebaceous Glands Neil Kitson, E.W. Van Lennep and J.A. Y o u n g * Departments of Physiology, and of Histology and Embryology, University of Sydney, Sydney,Australia Summary. A n examination o f h u m a n sebaceous glands by transmission electron m i c r o s c o p y has revealed the presence o f gap junctions. The junctions are f o u n d in a b u n d a n c e between differentiating cells, and annular forms are also seen. The possible significance o f this new finding is briefly discussed.
Key words: Sebaceous gland - H u m a n - G a p junctions.
Introduction Several authors have c o m m e n t e d on intercellular connections in h u m a n sebaceous glands (Charles, 1960; Hibbs, 1962; Ellis and Henrikson, 1963; Ellis, 1967; Fujita et al., 1972). While desmosomes have been reported in all these investigations, only two authors have published micrographs o f m e m b r a n e s in close apposition. Ellis (1967) c o m m e n t e d on the existence o f an "... occasional tight junction or nexus" and Bell (1974) noted "double walled structures" which she speculated were " p r o b a b l y specialized junctions". In our study o f h u m a n sebaceous glands, we have f o u n d m e m b r a n e s in close apposition having the characteristic appearance and dimensions o f gap junctions. These structures have been f o u n d in every gland so far examined.
Materials and Methods Tissue
Normal human skin was retrieved from ENT surgery in accordance with a protocol approved by the Medical Ethics Committees of the University of Sydney, and the Royal Prince Alfred Hospital, Sydney. Send offprint requests to: Dr. N. Kitson, Dept. of Physiology, University of Sydney, NSW 2006, Australia * We thank the ENT consultant staff of Royal Prince Alfred Hospital, Sydney, for making tissue available. The co-operation of the director, Dr. D.J.H. Cockayne, and staff of the University of Sydney Electron Microscope Unit is gratefully acknowledged. Mr. E. Foster provided excellent assistance with the photography. - The project was supported by the Consolidated Medical Research Funds, University of Sydney, and the National Health and Medical Research Council of Australia. One of us (N.K.) was the recipient of the Phyllis Anderson Medical Research Fellowship awarded by the Faculty of Medicine, University of Sydney
0302 766X/78/0190/0115/$01.40
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Samples were obtained in all cases from patients requiring post-auricular incisions.A strip of skin 1 mm wide was removed from the wound edge. This procedure had no influenceon the size or location of the initial incision.
Thin Sections
Excised skin was immediatelyimmersedin 2.5 % glutaraldehyde buffered with 0.1 M sodium cacodylate to pH 7.2. After 24 h primary fixation, samples were washed twice in buffer, post-fixed in cacodylatebuffered OsO~ for 2 h, block-stained for 45 min with 0.5 % uranyl acetate, rapidly dehydrated in an acetone series to a 50/50 mixture of Spurr's resin and acetone, and left overnight. Infiltration was accomplished over 48 h in two changes of 100 % resin, and the blocks were then embedded. Grey-to-silversectionswere cut on a Reichert OMU-3 ultramicrotome and mounted on naked, 400 mesh, copper grids. After staining with uranyl acetate and Reynolds' lead (Reynolds,1963), sectionswere viewed in a Philips 201 electron microscope operating at 60 KV with a 50 mcm objective aperture.
Lanthanum
Some tissue blocks were processed with lanthanum hydroxide present in all solutions up to the dehydration sequence after the method of Revel and Karnovsky (1967). Otherwise, the tissue was handled as for thin sections.
Results G a p j u n c t i o n s could be seen at lower powers (Figs. 1-3) as dense bands. A t higher powers (Figs. 4, 6), the characteristic "seven layered" structure ( B r i g h t m a n a n d Reese, 1969) became apparent, a n d the gap was discernible as were periodic crossstriations. The j u n c t i o n widths m e a s u r e d between 1 6 . 5 - 1 8 . 0 n m in r a n d o m m e a s u r e m e n t s with a n optical micrometer. The gap widths were a b o u t 2 - 4 nm, a n d l a n t h a n u m infiltrated j u n c t i o n s (Fig. 5) showed a dense central b a n d 4 - 6 n m wide. T h e l a n t h a n u m could n o t be distinguished from the electron-dense outer leaflets of the a p p o s i n g m e m b r a n e s . These results are in general agreement with those of other a u t h o r s (Staehelin, 1974). The hexagonal p a t t e r n usually seen in oblique sections of l a n t h a n u m - t r e a t e d j u n c t i o n s (Revel a n d K a r n o v s k y , 1967) has n o t been observed in our preparations. A n n u l a r j u n c t i o n s were occasionally seen (Fig. 6). The area enclosed by the j u n c t i o n always a p p e a r e d less electron-dense t h a n the s u r r o u n d i n g cytoplasm, b u t n o recognizable organelles were seen within the j u n c t i o n s . We have n o t attempted to determine whether or n o t a c o n n e c t i o n exists between these structures a n d the cytoplasmic m e m b r a n e , b u t so far a n n u l a r j u n c t i o n s have n o t been seen in l a n t h a n u m - t r e a t e d tissue. A l t h o u g h a q u a n t i t a t i v e analysis c a n n o t be offered, a n impression has been gained that gap j u n c t i o n s are f o u n d almost entirely between differentiating sebaceous cells, a n d n o t between undifferentiated or fully differentiated cells.
G a p Junctions in H u m a n Sebaceous Glands
117
Fig. 1. Sebaceous cells from a 13 year old male ( x 15,000). Arrows show two gap junctions in apposing membranes of differentiating cells. L lipid Fig. 2. Sebaceous cells from a 13 year old male ( x 30,000). A n extensive junction is seen between the arrows. L lipid Fig.3. Sebaceous cells from a 13 year old male ( x 25,000). Arrow marks small junction
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Fig.4. Sebaceous cells from a 13 year old male ( x 236,000). The gap is visible as are periodic cross striations Fig. 5. Sebaceous cells from a 44 year old male ( x 261,000). L a n t h a n u m has infiltrated the gap and obscured the electron dense outer leaflets. Periodic electron lucent areas are faintly visible Fig. 6. Sebaceous cells from a 13 year old female ( x 79,000). A n annular junction is shown with enclosed area appearing less electron dense than surrounding cytoplasm. Inset ( • 299,000) shows gap in same junction
Gap Junctions in Human SebaceousGlands
119
This is the first report of gap junctions in sebaceous glands although the structures previously reported by Ellis (1967) and Bell (1974) were undoubtedly also gap junctions. Such junctions have been observed in wool follicles (Orwin et al., 1973), rat oral mucosa (Shimono and Clementi, 1976) and human epidermis (Wolff and Schreiner, 1968; Hashimoto, 1971; Vreeswijk et al., 1977). Cavoto and Flaxman (1972) found gap junctions between epidermal cells in culture, and also established the existence of low-resistance electrical connections between such cells. Gap junction subunits are thought to be intercellular channels (Revel and Karnovsky, 1967; Brightman and Reese, 1969; Peracchia, 1973; Caspar et al., 1977; Makowski et al., 1977) which allow electrical coupling (Loewenstein and Kanno, 1964; Rose, 1971; Ito et al., 1974) and metabolic co-operation (Gilula et al., 1972; Azarnia et al., 1974; Pitts and Simms, 1977). Electrical coupling has been observed in several exocrine glands that respond to autonomic stimulation (Ginsborg et al., 1974; Petersen and Ueda, 1976), but the role of membrane conductance changes in secretion is not clear. Lawrence et al. (1977) have evidence to suggest that intra cellular messengers are transmitted through gap junctions; membrane conductance changes in coupled cells may conceivably be secondary to such phenomena. In the case of sebaceous glands, secretion involves cell lysis (Palay, 1958) after several days' differentiation (Downing et al., 1975). In addition, the glands are said not to be innervated (Montagna, 1963), and thus close autonomic control of secretion by regulation of membrane conductance is unlikely; exchange of chemical information through the junctions is more probable. However, Dugan (1974) and F6rster (1975) have reported autonomic nerves innervating sebaceous gland basal cells respectively, in rat and mouse, and the question must remain open. "Annular" gap junctions have been observed in other tissues (Garant et al., 1972; Merk et al., 1973; Orwin et al., 1973; Bjersing and Cajander, 1974; Albertini et al., 1975), in tissue culture (Perissel et al., 1976), in tumour (Letourneau et al., 1975), and in epidermis (Vreeswij k et al., 1977). By serial sectioning, Vreeswijk et al. showed that the junction limited vesicles so studied were not connected to cytoplasmic membrane. In other material however, Merk et al. (1973), Letourneau et al. (1975), and Perissel et al. (1976) showed that lanthanum infiltrated some annular junctions, implying continuation with the intercellular space. Van Lennep and Madden (1965) observed similar membrane-enclosed vesicles in human corpus luteum, and although they described the membranes as"quintuple layered" as found in the zonula occludens, the structures are more likely gap junctions of the annular type. These authors interpreted their findings as a "pinching off' of cytoplasmic membrane, followed by destruction of the enclosed material. Albertini et al. (1975) speculated that such "interiorization" of gap junctions may be a method of removing the restricting influence of these connections on cell movements. Thus, a cell might repeatedly make and break connections with adjacent cells while moving through an epithelium. While van Lennep and Madden, and Albertini et al. were referring to ovarian granulosa cells, the same hypothesis could easily be extended to epidermal and sebaceous cells. Interestingly, the frequency of annular junctions found in estrogen-dependent renal adenocarcinoma appears to decrease when estrogen is withdrawn (Letourneau et al., 1975).
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The finding of gap junctions between sebaceous cells is of interest, although perhaps not unexpected given the prevalence of these connections in epithelia generally. The concept, however, that intercellular communication may play a role in the regulation of this form of holocrine secretion is attractive. It remains to be established that the characteristic hexagonal subunit pattern exists between the membranes of such junctions, and that low resistance pathways are present between cells so connected. References Albertini, D.F., Fawcett, D.W., Olds, P.J.: Morphological variations in gap junctions of ovarian granulosa cells. Tissue and Cell 7, 389-405 (1975) Azarnia, R., Larsen, W.J., Loewenstein, W.R.: The membrane junctions in communicating and noncommunicating cells, their hybrids, and segregants. Proc. nat. Acad. Sci. (Wash.) 71, 880-884 (1974) Bell, M.: A comparative study of the ultrastructure of the sebaceous glands of man and other primates. J. invest. Dermatol. 62, 132-143 (1974) Bjersing, L., Cajander, S.: Ovulation and the mechanism of follicle rupture. IV. Ultrastructure of membrana granulosa of rabbit Graafian follicles prior to induced ovulation. Cell Tiss. Res. 153,1-14 (1974) Brightman, M.W., Reese, T.S.: Junctions between intimately apposed cell membranes in the vertebrate brain. J. Cell Biol. 40, 648~77 (1969) Caspar, D.L.D., Goodenough, D.A., Makowski, L., Phillips, W.C.: Gap junction structures. I. Correlated electron microscopy and X-ray diffraction. J. Cell Biol. 74, 605-628 (1977) Cavoto, F.V., Flaxman, B.A.: Communication between normal human epidermal cells in vitro. J. invest. Dermatol. 59, 370-374 (1972) Charles, A.: Electron microscopic observations of the human sebaceous gland. J. invest. Dermatol. 35, 31-36 (1960) Downing, D.T., Strauss, J.S., Ramasastry, P., Abel, M., Lees, C.W., Pochi, P.E.: Measurement of the time between synthesis and surface excretion of sebaceous lipids in sheep and man. J. invest. Dermatol. 64, 215-219 (1975) Dugan, K.H.: Ultrastructural observations of possible nerve endings in rat sebaceous glands. Cell Tiss. Res. 150, 333-348 (1974) Ellis, R.A.: Eccrine, sebaceous, and apocrine glands. In: Ultrastructure of Normal and Abnormal Skin. 1st ed. (A.S. Zelickson, ed.), pp. 132-162. London: Henry Kimpton 1967 Ellis, R.A., Henrikson, R.C.: The ultrastructure of the sebaceous glands of man. In: Advances in Biology of Skin, Vol. IV (W. Montagna, R.A. Ellis and A.F. Silver, eds.), pp. 94-109. Oxford: Pergamon Press 1963 F6rster, F2., Heine, H., Schaeg, G.: Histophysiology of the vegetative peripheral nervous system of skin. Arch. Dermatol. Res. 254, 295-302 (1975) Fujita, H., Asagami, C., Murata, S., Murozumi, S.: Ultrastructural study of embryonic sebaceous cells, especially of their sebum droplet formation. Acta derm.-venerol. (Stockh.) 52, 99-115 (1972) Garant, P.R.: The demonstration of complex gap junctions between the cells of the enamel organ with lanthanum nitrate. J. Ultrastruct. Res. 40, 333-348 (1972) Gilula, N.B., Reeves, O.R., Steinbach, A.: Metabolic coupling, ionic coupling, and cell contacts. Nature (Lond.) 235, 262-265 (1972) Ginsborg, B.L., House, C.R., Silinsky, E.M.: Conductance changes associated with the secretory potential in the cockroach salivary gland. J. Physiol. (Lond.) 236, 723-731 (1974) Hashimoto, K.: Intercellular spaces of the human epidermis as demonstrated with lanthanum. J. invest. Dermatol. 57, 17-31 (1971) Hibbs, R.G. : Electron microscopy of human axillary sebaceous glands. J. invest. Dermatol. 38, 329-336 (1962) Ito, S., Sato, E., Loewenstein, W.R.: Studies on the formation of a permeable cell membrane junction. II. Evolving junctional conductance and junctional insulation. J. Membrane Biol. 19, 339-355 (1974)
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Lawrence, T.S., Beers, W.H., Gilula, N.B.: Hormonal stimulation and cell communication in cocultures. J. Cell Biol. 75, 63a (1977) Letourneau, R.J., Li, J.J., Rosen, S., Villee, C.A.: Junctional specialization in estrogen-induced renal adenocarcinomas of the golden hamster. Cancer Res. 35, 6-10 (1975) Loewenstein, W.R., Kanno, Y.: Studies on an epithelial cell junction. I. Modifications of surface membrane permeability. J. Cell Biol. 22, 565-586 (1964) Makowksi, L., Caspar, D.L.D., Phillips, W.C., Goodenough, D.A.: Gap junction structures. II. Analysis of the X-ray diffraction data. J. Cell Biol. 74, 629~545 (1977) Merk, F.B., Albright, J.T., Botticelli, C.R. : The fine structure of granulosa cell nexuses in rat ovarian follicles. Anat. Rec. 175, 107-125 (1973) Montagna, W.: The sebaceous glands in man. In:Advances in Biology of Skin. Vol. IV. The Sebaceous Glands, (W. Montagna, R.A. Ellis, and A.F. Silver, eds.), pp. 19-35. Oxford: Pergamon Press 1963 Orwin, D.F.G., Thomson, R.W., Flower, N.E.: Plasma membrane differentiations of keratinizing cell wool follicles. I. Gap junctions. J. Ultrastruct. Res. 45, 1-14 (1973) Palay, S.L.: The morphology of secretion. In: Frontiers in Cytology. (S.L. Palay, ed.), pp. 305-342. New Haven: Yale University Press 1958 Peracchia, C.: Low resistance junctions in crayfish. I. Two arrays of globules in junctional membranes. J. Cell Biol. 57, 54-76 (1973) Perissel, B., Charbonnt, F., Chessebeuf, M., Malet, P.: Differentiation of the plasma membrane of hepatic cells in monolayer cultures. Cell Tiss. Res. 171, 157-173 (1976) Petersen, O.H., Ueda, N.: Pancreatic acinar cells: the role of calcium in stimulus-secretion coupling. J. Physiol. (Lond.) 254, 583-606 (1976) Pitts, J.D., Simms, J.W.: Permeability of junctions between animal cells. Exp. Cell Res. 104, 153-163 (1977) Revel, J.P., Karnovsky, M.I. : Hexagonal array of subunits in intercellular junctions of the mouse heart and liver. J. Cell Biol. 33, C7-C17 (1967) Reynolds, E.S.: The use of lead citrate at high pH as an electron-opaque stain in electron microscopy. J. Cell Biol. 17, 208-213 (1963) Rose, B.: Intercellular communication and some structural aspects of membrane junctions in a simple cell system. J. Membrane Biol. 5, 1-19 (1971) Shimono, M., Clementi, F.: Intercellular connections of oral epithelium. I. Studies with freeze-fracture and tracing methods of normal rat keratinized oral epithelium. J. Ultrastruct. Res. 56, 121-136 (1976) Staehelin, A.: Structure and function of intercellular junctions. Int. Rev. Cytol. 39, 191-283 (1974) Van Lennep, E.W., Madden, L.M.: Electron microscope observations on the involution of the human corpus luteum of menstruation. Z. Zellforsch. 66, 365-380 (1965) Vreeswijk, J., Leene, W., Kalsbeek, G.L.: Early host cell - Molluscum contagiosum virus interactions. II. Viral interactions with the basal epidermal cells. J. invest. Dermatol. 69, 249-256 (1977) Wolff, K., Schreiner, E.: An electron microscopic study on the extraneous coat of keratinocytes and the intercellular space of the epidermis. J. invest. Dermatol. 51, 418430 (1968)
Accepted February 9, 1978
