The Roles of Indian Hedgehog Signaling in TMJ Formation.

The temporomandibular joint (TMJ) is an intricate structure composed of the mandibular condyle, articular disc, and glenoid fossa in the temporal bone. Apical condylar cartilage is classified as a secondary cartilage, is fibrocartilaginous in nature, and is structurally distinct from growth plate and articular cartilage in long bones. Condylar cartilage is organized in distinct cellular layers that include a superficial layer that produces lubricants, a polymorphic/progenitor layer that contains stem/progenitor cells, and underlying layers of flattened and hypertrophic chondrocytes. Uniquely, progenitor cells reside near the articular surface, proliferate, undergo chondrogenesis, and mature into hypertrophic chondrocytes. During the past decades, there has been a growing interest in the molecular mechanisms by which the TMJ develops and acquires its unique structural and functional features. Indian hedgehog (Ihh), which regulates skeletal development including synovial joint formation, also plays pivotal roles in TMJ development and postnatal maintenance. This review provides a description of the many important recent advances in Hedgehog (Hh) signaling in TMJ biology. These include studies that used conventional approaches and those that analyzed the phenotype of tissue-specific mouse mutants lacking Ihh or associated molecules. The recent advances in understanding the molecular mechanism regulating TMJ development are impressive and these findings will have major implications for future translational medicine tools to repair and regenerate TMJ congenital anomalies and acquired diseases, such as degenerative damage in TMJ osteoarthritic conditions.

Part I: Development and Physiology of the Temporomandibular Joint.

PURPOSE OF REVIEW: Investigate the developmental physiology of the temporomandibular joint (TMJ), a unique articulation between the cranium and the mandible. RECENT FINDINGS: Principal regulatory factors for TMJ and disc development are Indian hedgehog (IHH) and bone morphogenetic protein (BMP-2). The mechanism is closely associated with ear morphogenesis. Secondary condylar cartilage emerges as a subperiosteal blastema on the medial surface of the posterior mandible. The condylar articular surface is immunoreactive for tenascin-C, so it is a modified fibrous periosteum with an underlying proliferative zone (cambrium layer) that differentiates into fibrocartilage. The latter cushions high loads and subsequently produces endochondral bone. The TMJ is a heavily loaded joint with three cushioning layers of fibrocartilage in the disc, as well as in subarticular zones in the fossa and mandibular condyle. The periosteal articular surface produces fibrocartilage to resist heavy loads, and has unique healing and adaptive properties for maintaining life support functions under adverse environmental conditions.

Spry1 and spry2 are essential for development of the temporomandibular joint.

The temporomandibular joint (TMJ) is a specialized synovial joint essential for the function of the mammalian jaw. The main components of the TMJ are the mandibular condyle, the glenoid fossa of the temporal bone, and a fibrocartilagenous disc interposed between them. The genetic program for the development of the TMJ remains poorly understood. Here we show the crucial role of sprouty (Spry) genes in TMJ development. Sprouty genes encode intracellular inhibitors of receptor tyrosine kinase (RTK) signaling pathways, including those triggered by fibroblast growth factors (Fgfs). Using in situ hybridization, we show that Spry1 and Spry2 are highly expressed in muscles attached to the TMJ, including the lateral pterygoid and temporalis muscles. The combined inactivation of Spry1 and Spry2 results in overgrowth of these muscles, which disrupts normal development of the glenoid fossa. Remarkably, condyle and disc formation are not affected in these mutants, demonstrating that the glenoid fossa is not required for development of these structures. Our findings demonstrate the importance of regulated RTK signaling during TMJ development and suggest multiple skeletal origins for the fossa. Notably, our work provides the evidence that the TMJ condyle and disc develop independently of the mandibular fossa.

Immunohistochemical expression of types I and III collagen antibodies in the temporomandibular joint disc of human foetuses.

The objective was to study the morphology of the articular disc and analyse the immunohistochemical expression of types I and III collagen markers in the temporomandibular joint (TMJ) disc of human foetuses of different gestational ages. Twenty TMJ from human foetuses supplied by Universidade Federal de Uberaba with gestational ages from 17 to 24 weeks were studied. The gestational age of the foetuses was determined by measuring the crown-rump (CR) length. Macroscopically, the foetuses were fixed in 10% formalin solution and dissected by removing the skin and subcutaneous tissue and exposing the deep structures. Immunohistochemical markers of type I and III were used to characterize the existence of collagen fibres. Analysis of the immunohistochemical markers of types I and III collagen revealed the presence of heterotypical fibril networks.

Human temporomandibular joint disc: anatomy and measurements in prenatal development.

The objective of this study was to determine morphological characteristics and measurements of the temporomandibular joint (TMJ) disc in human fetuses between 16 and 20 weeks of intrauterine life, and correlate it with oral-facial neuro-muscular maturing. Scanner images were used to record the length of the disc (D) and the thickness of its anterior middle and posterior bands in TMJ anteroposterior vertical sections from human fetuses of 16, 18 and 20 weeks of intrauterine life (WIL). Mean disc length was 1.98 mm, 2.69 mm and 2.90 mm at 16, 18 and 20 WIL respectively, and measurements differed significantly between those ages. The thicknesses of the anterior, middle and posterior bands also differed significantly. The results give normal morphological data for D between 16 and 20 WIL. TMJ anatomy and measurements appear to be related and agree with the neuro-muscular maturation time at which sucking and swallowing reflexes begin before birth. It is known that these functions, as well as the neuro-muscular capacity to perform prenatal mandibular movements (opening and closing), begin at 14 to 15 weeks of prenatal development and are fully attained at about 20 weeks of development. Knowledge of this reference pattern may be of major importance to future research, for assessing jaw biomechanics and detecting alterations of TMJ and prenatal development of a vital human function - suckling in preterm infants.

Human temporomandibular joint disc: anatomy and measurements in prenatal development

El objetivo de este estudio fue determinar las características morfológicas y dimensionales del disco de la articulación temporomandibular (ATM) de fetos humanos entre las 16 y 20 semanas de vida intrauterina y correlacionarla con la maducación neuro-muscular bucofacial. Mediante un analizador de imágenes se registró la longitud sagital, el espesor de las bandas anterior, media y posterior del disco (D) en corte vertical anteroposterior de ATM de fetos humanos de 16, 18 y 20 semanas de vida intrauterina (SVI). La longitud promedio del disco fue de 1.98mm, 2.69mm y 2.90mm respectivamente, encontrándose diferencias dimensionales significativas entre aquellas edades. También los espesores anterior, intermedio y posterior de las bandas mostraron diferencias significativas entre los D de las edades estudiadas. Los resultados proporcionaron datos morfológicos normales del D entre las 16 y 20 SVI. Los parámetros anatómicos y dimensionales de la ATM encontrados estarían relacionados y coinciden con elt iempo de maduración neuromuscular en el cual se inician los actos reflejos de succión y deglución previa al nacimiento. Como es conocido, aquellas funciones se inician entre las 14 o 15 semanas de gestación, casí como la capacidad neuromuscular para realizar los movimientos mandibulares prenatales (apertura y cierre) que se alcanzarían alrededor de las 20 semanas de desarrollo. El conocimiento de este patrón de referencia puede ser de gran importancia para futuras investigaciones, para valorar la biomecánica mandibular y permitir reconocer alteraciones de la ATM y del desarrollo prenatal en una de las funciones vitales del ser humano como es la lactancia en reción nacidos prematuros.

A direct anatomical study of the morphology and functionality of disco-malleolar and anterior malleolar ligaments / Estudio anatómico directo de la morfología y funcionalidad de los ligamentos disco-maleolar y maleolar anterior

The disco-malleolar and anterior malleolar ligaments are common to the middle ear, the temporomandibular joint and the jaw, all of them sharing a common embryological origin; these ligaments' morphometric and functional aspects were studied in 23 temporal bones. Experimental design: The epitympanum roof and the temporomandibular joint roof were elevated by micro-dissection of the middle cranial fossa, exposing both disco-malleolar and anterior malleolar ligaments and their association with the malleus, the temporomandibular joint disc and the mandibular lingula. Principal observations: Both ligaments start in the malleus and take anterior and divergent routes towards the temporomandibular joint and lingula, passing through Huguier's canal. The disco-malleolar and anterior malleolar ligaments' mean lengths were 6.88 mm (SD 0.81) and 4.22mm (SD 1.17), respectively, no statistically significant difference being revealed between the sides. Temporomandibular joint disc traction was applied for verifying malleus mobility; malleus movement was observed when applying disco-malleolar traction in 30.5% of the samples. Correlation was observed between malleus movement and disco-malleolar length (R2=-0.499, p<0.05). Both ligaments common to the stomatognathic system and middle ear were present in all specimens. Conclusions: There was an anatomical and functional relationship between human TMJ and the middle ear.

Los ligamentos disco-maleolar y maleolar anterior son comunes en el oído medio, la articulación temporomandibular y la mandíbula, todas ellos comparten un origen embriológico común. Los aspectos morfométricos y funcionales de estos ligamentos fueron estudiados en 23 huesos temporales. Diseño experimental: el techo del epitímpano y el techo de la articulación temporomandibular fue elevada mediante micro-disección de la fosa craneal media, exponiendo ambos ligamentos disco-maleolar y maleolar anterior y su asociación con el maleus, el disco de la articulación temporomandibular y língula mandibular. Principales observaciones: Ambos ligamentos comienzan en el maleus y toman una ruta anterior y divergente hacia la articulación temporomandibular y língula, pasando a través del canal de Huguier. Las longitudes medias del ligamento disco-maleolar y maleolar anterior fueron 6,88 mm (DS 0,81) y 4,2mm (DS 1,17), respectivamente, no fueron reveladas diferencias estadísticamente significativas entre las partes. La tracción del disco de la articulación temporomandibular fue aplicada para verificar la movilidad del maleus; el movimiento del maleus fue observado cuando se aplicó tracción del ligamento disco-maleolar en el 30,5% de las muestras. Se observó correlación entre el movimiento del maleus y la longitud disco-maleolar (R2 = -0,499, p <0,05). Ambos ligamentos comunes al sistema estomatognático y el oído medio estaban presentes en todos los especímenes. Conclusiones: Existe una relación anatómica y funcional entre la ATM humana y el oído medio.

Immunohistochemical expression of collagen type IV antibody in the articular disc of the temporomandibular joint of human fetuses.

The objective of this paper was to study the morphology of the articular disc and analyze the immunohistochemical expression of the marker of type IV collagen in the articular disc of the temporomandibular joint (TMJ) of human fetuses of different gestational ages. Twenty TMJ from human fetuses aging from 21 to 24 weeks of intrauterine life were studied. The TMJ were supplied by the Federal University of Uberaba. The ages of the fetuses were determined by measuring the crown-rump length (CRL). Macroscopically, the fetuses were fixed in a formalin solution at 10% and dissected by removing the skin and the subcutaneous tissue, exposing the deep structures. An immunohistochemical marker of type IV collagen was used in order to characterize the presence of blood vessels in the central region of the temporomandibular joint disc. Analysis of the immunohistochemical marker of type IV collagen showed the presence of blood vessels in the central region of the temporomandibular disc in human fetuses.

Human temporomandibular joint morphogenesis.

Temporomandibular joint morphogenesis was studied. Ranging in age of fetuses examined was from 6 to14 weeks' gestation. Our results showed the condyle so first element that appear between 6 degrees and 8 degrees week (condylar blastema). After a week appear temporal elements. Disk appear at the same time of glenoid blastema and it reaches an advanced differentation before of the condyle and temporal element, so these don't effect machanical compression on mesenchyma where we find the disk. So we think that the disk result of genetic expression and it isn't the result of mechanical compression. The inferior joint cavity appear to 12 week. The superior joint cavity appear to 13-14 week. In conclusion, the appearance of the condyle is the first event during TMJ morphogenesis, with its initial bud, in form of a mesenchymal thickening, becoming detectable between the sixth and eight week of development, when all the large joints of the limbs are already well defined.

Morphology of the lateral pterygoid muscle associated to the mandibular condyle in the human prenatal stage.

The lateral pterygoid muscle (LPM) inserts at the condyle and the articular disc and plays a central role in mandibular movement via the Temporomandibular Articular Complex. The aim of this study was to examine the association between the morphology of LPM muscular fascicles and the degree of mineralization of the mandibular condyle in the prenatal stage employing structural, ultrastructural and microanalytical evaluation. Sixteen human fetuses at 11-37 weeks of gestation, with no apparent pathology and resulting from spontaneous abortions, were included in the study. Samples from lateral pterygoid muscle and the mandibular condyle were processed for light microscopy and electron microscopy and microanalysis. Desmin immunolabeling (dilution 1: 25 Dako) and alpha sarcomeric actin immunolabeling (dilution 1:50 Dako) employing the avidin-biotin system were used in paraffin embedded samples. Contralateral samples were examine by transmission electron microscopy. Four condyles (at 17-21 weeks of gestation) were used to measure the relative content of calcium and phosphorous employing the X-ray diffraction microanalytical technique. At 11-16 weeks of gestation, the LPM was composed of secondary myotubes associated to satellite cells and nerve fibers. At 18 weeks, the muscle exhibited multiple compact fascicles and the condyle showed a thin, external, subperiostal mineralized layer with few central bone spicules. At 20 weeks, at the site of insertion of the LPM, the bone trabeculae of the condyle contained an electrondense matrix with abundant mineralization nuclei. At 17-21 weeks of gestation no significant variations in the contents of phosphorous and calcium were observed. At 24 weeks, transmission electron calcium and microscopy studies revealed a marked increase in the functional units of the muscle fascicles. Also, at this age muscle fibers exhibited differences in the expression of desmin and alpha sarcomeric actin. At 37 weeks the muscle became multipennate in appearance, exhibiting a more complex organization than younger fetuses. Alpha sarcomeric actin labeling became light with age. This results suggest that between 16 and 22 weeks of gestation the differentiation and maturation process of the muscle fibers precedes and prevails over the development and mineralization process from mandibular condyle. The rudimentary performance of the prenatal LPM would be one of the factors that regulate the process of ossification at the level of the mandibular condyle. The rate of ossification would increase starting from 22 of gestation week.

Distribution of mucopolysaccharides and glycoproteins in the articular discs of temporomandibular joints in human fetuses.

AIMS: To evaluate the distribution of mucopolysaccharides and glycoproteins in the articular discs of temporomandibular joints (TMJs) in human fetuses at different stages of development in order to test the hypothesis that the development and histological maturation of the articular disc has already begun by the 12th week of gestation. METHODS: Eighteen human fetuses at gestational ages 12, 14, and 16 weeks were used (6 fetuses of each age). Sections (6 microm wide) of the articular discs were stained with trichromic stain for collagen fibers, Mayer's mucicarmine for mucopolysaccharides, and Schiff's periodic acid reaction for glycoproteins. The densities of the stained zones were measured by means of Image J software. The nonparametric Kruskal-Wallis test was used to evaluate the differences among stained zones in the 3 fetus groups. RESULTS: TMJ tissues of four of the six 12-week-old fetuses were stained positive for collagen fibers, mucopolysaccharides, and glycoproteins. In these fetuses the stain was localized to the articular posterior area and was denser in that area than in the middle and anterior areas. The stained areas in the 14-week-old fetuses were distributed throughout the articular discs, with isolated clear unstained areas. The stained areas of all the articular discs of the 16-week-old fetuses were more compact than those of the 12-and 14-week-old fetuses. In all the fetuses examined, the collagen fibers along the articular disc had a wavy appearance. The fossa of the temporal bone was observed in all the fetuses as a straight structure that was similar in the 3 fetus groups. The densities (mean +/- SD) of the stained zones were 38.36% +/- 3.39%, 59.5% +/- 1.56%, and 94.04% +/- 2.04% for 12, 14, and 16 weeks of gestation, respectively; these densities were significantly different (chi2 = 15.16; df = 2, P < .001). CONCLUSION: This study indicates that mucopolysaccharides and glycoproteins as well as collagen fibers are present at 12, 14, and 16 weeks of gestation. This suggests that the histological maturation of the articular disc has already begun at the 12th week and is complete by the 16th week of gestation.

Histochemistry of the foetal human temporomandibular joint articular disc.

The human temporomandibular joint (TMJ) develops from mesenchymal cells that form condensations appearing as condylar and temporal blastema which give rise to the respective anterior and posterior regions of the TMJ articular disc. Previous reports have shown the foetal disc to be avascular, with a high content of organized collagen fibres and a lesser content of elastic fibres. In this study, the articular discs from TMJs of a human foetus at age 22 weeks were evaluated. At this stage of intrauterine (i.u.) development, the disc was found to be a highly cellular, biconcave structure with a dense arrangement of collagen fibres. Cell density was not uniform, with increased density in the intermediate band relative to the anterior and posterior bands. In contrast to earlier reports, capillaries containing red blood cells were observed along the inferior surface of the disc. Immunohistochemical staining for proteoglycans and glycosaminoglycans (GAGS) revealed abundant chondroitin sulphate proteoglycan (CSPG) and hyaluronic acid in the disc while relatively little amounts of dermatan sulphate proteoglycan II (DSPGII) were found. No keratin sulphate proteoglycan (KSPG) was detectable. Foetal human TMJ articular discs at this age were found to have morphology and regional characteristics similar to adult discs.

Early foetal development of the articular disc in the human temporomandibular joint.

In foetuses of 9 and 10 weeks the articular disc presents a more cellular structure with bands of connective tissue fibres. It is connected with the articular capsule and lateral pterygoid muscle. During weeks 11 and 12 there is an increase in collagenous fibres and fusiform cells are located mainly close to the surface of the disc.

Expression of heat shock protein 27 (HSP27) in human temporomandibular joint discs of patients with internal derangement.

Heat shock proteins (HSPs) are believed to represent a cellular stress response mechanism that protects intracellular proteins from damaging events. Some studies have demonstrated an enhanced expression of large-molecular-weight HSPs in diseased systematic joints. Small heat shock proteins, and among these HSP27, have been studied to a lesser extent. HSP27 has cytoprotective and biosynthetic functions within chondrocytes, and it is an estrogen-associated protein that is under hormonal modulation. To improve understanding at a molecular level of the pathophysiology of certain temporomandibular joint disorders, the authors carried out this immunohistochemical study to assess the presence of HSP27 in human TMJ discs. Twelve adult human TMJ discs (10 diseased and 2 healthy discs) and 5 TMJ fetal human discs were used in this study. Adult discs and TMJ tissues of human fetuses were fixed in 10% buffered formalin. Samples were then processed for histologic examination. Sections were immunohistochemically stained using the streptavidin-biotin detection method. No reaction product for HSP27 in the discs of fetuses was noted. HSP27 was weakly expressed in normal TMJ discs and highly up-regulated in discs of patients showing new vessel formation and chondroid metaplasia. Any correlation between gender and HSP27 was found in the sample, being the up-regulation of HSP27 related mostly to major histopathological changes. This different pattern of HSP27 immunostaining in human TMJ discs detected in the authors' specimens suggests that the expression of this small HSP is functionally modulated. In fact HSP27 up-regulates in internal derangement specimens with major histopathological changes; on the other hand, it is not expressed or only weakly expressed in TMJ discs of fetuses and normal TMJ discs.

Development of the human temporomandibular joint.

A great deal of research has been published on the development of the human temporomandibularjoint (TMJ). However, there is some discordance about its morphological timing. The most controversial aspects concern the moment of the initial organization of the condyle and the squamous part of the temporal bone, the articular disc and capsule and also the cavitation and onset of condylar chondrogenesis. Serial sections of 70 human specimens between weeks 7 and 17 of development were studied by optical microscopy (25 embryos and 45 fetuses). All specimens were obtained from collections of the Institute of Embryology of the Complutense University of Madrid and the Department of Morphological Sciences of the University of Granada. Three phases in the development of the TMJ were identified. The first is the blastematic stage (weeks 7-8 of development), which corresponds with the onset of the organization of the condyle and the articular disc and capsule. During week 8 intramembranous ossification of the temporal squamous bone begins. The second stage is the cavitation stage (weeks 9-11 of development), corresponding to the initial formation of the inferior joint cavity (week 9) and the start condylar chondrogenesis. Week 11 marks the initiation of organization of the superior joint cavity. And the third stage is the maturation stage (after week 12 of development). This work establishes three phases in TMJ development: 1) the blastematic stage (weeks 7-8 of development); 2) the cavitation stage (weeks 9-11 of development); and 3) the maturation stage (after week 12 of development). This study identifies the critical period of TMJ morphogenesis as occurring between weeks 7 and 11 of development.

Development of the human temporomandibular joint. Computer-aided 3D-reconstructions.

Computer-aided graphical three-dimensional reconstructions of histological serial sections of 12 human embryos and fetuses (25-250 mm Crown-rump length (CRL)) were used to trace the prenatal development of the elements of the human temporomandibular joint. The primordia of the condylar and coronoid processes could be identified as two bony peaks at the dorsal ends of the mandible at the stage of 25 mm CRL. The primordium of the temporal bone already existed at the stage of 37 mm CRL. The bone was apparent with a convex contour towards the condyle. The glenoid fossa was not yet visible. At 65 mm CRL, the osseous glenoid fossa could be distinguished at the enlarged temporal bone formation. The glenoid fossa developed posteriorly and medially from the condyle and extended in cranial and anterior direction. The glenoid fossa had various contours, changing from flat and slightly convex (65 mm CRL) to concave (250 mm CRL) with an articular tubercle. The distance between fossa and condyle increased proportionally. The lower joint cavity appeared at an earlier stage (65 mm CRL) than the upper joint cavity (70 mm CRL). Both cavities started development as isolated compartments fusing later on. The upper joint cavity followed the contour of the fossa, whereas the lower joint cavity followed the form of the condyle. The biconcave shape of the articular disc as well as the attachment of the lateral pterygoid muscle could be observed very early (70 mm CRL).

The vascular relationship between the temporomandibular joint and the middle ear in the human fetus.

PURPOSE: The aim of this work was to clarify the vascular relationships between the middle ear and the temporomandibular joint region during human fetal development. MATERIALS AND METHODS: Light microscopic studies were done on 40 human fetuses from 72 mm crown-rump length (C-R) to 150 mm C-R, which were stained by various methods. Five human fetuses were dissected. Natural latex with industrial coloring was injected through the external carotid artery. All specimens were dissected bilaterally. RESULTS: The limits of the retroarticular region and the fetal tympanosquamosal fissure are shown. The anterior tympanic artery has a variable origin. In most cases, it originates from the maxillary artery; in other cases it originates from the superficial temporal artery or the bifurcation of the external carotid artery. On its way through the retroarticular region, it gives branches to the posterior part of the temporomandibular joint. It progresses along the most lateral part of the tympanosquamosal fissure, dividing into three branches that extend throughout the middle ear. A number of venous spaces in the retroarticular region that constitute the retrodiscal venous plexus. Small venous vessels along the fetal tympanosquamosal fissure accompany the anterior tympanic artery and drain into the retrodiscal venous plexus. CONCLUSIONS: During human fetal development, there is a wide connection across the tympanosquamosal fissure between the middle ear and the temporomandibular joint region. The anterior tympanic artery and its branches, as well as small venous vessels that are connected with the retrodiscal venous plexus, extend along the most lateral part of the fissure.

[Oto-mandibular ligaments: disco-mallear and malleo-mandibular ligaments]. / Les ligaments oto-mandibulaires: ligaments disco-malléaire et malléo-mandibulaire.

Phylogenesis, embryology and anatomy are emphasizing that two ligaments are found between the temporo-mandibular joint and the middle ear. These two oto-mandibular ligaments are named the disco-mallear ligament and the malleo-mandibular ligament. Originating from the first arch, these ligaments are not involved in the otologic manifestations of the temporo-mandibular joint syndrome. The disco-mallear ligament is a brake applied to the anterior excursion of the disc. When this disco-mallear ligament is stretched the disc can be displaced anteriorly breeding disc displacement, hypermobility and temporo-mandibular dislocation. The malleo-mandibular ligament is a remainder of the Meckel's cartilage: its role is not clear. When excessive forces are applying on the mandible, ossicles dislocation can be seen the forces being transmitted through it.

Changes in the chemical composition of the bovine temporomandibular joint disc with age.

The bovine temporomandibular joint disc is a fibrocartilaginous structure composed largely of collagen and proteoglycans. Little is known about changes in its composition accompanying growth and maturation. Discs were collected from immature foetuses (3-5 months), mature foetuses (6-8 months, adolescents (18 months), young adults (2-3 yr) and mature adults (over 4 yr), dissected free of fibrous attachments, and separated into outer and inner tissues. For the outer tissues the major findings were that: (1) water content in postnatal specimens was less than in prenatal specimens: (2) collagen content (relative to tissue dry weight) increased up to adolescence with little change thereafter; (3) total glycosaminoglycan, chondroitin sulphate and hyaluronic acid contents decreased during foetal development and then remained relatively constant, and (4) dermatan sulphate (the major glycosaminoglycan at all ages) decreased at maturity while keratan sulphate increased slightly. Results for the inner tissues were similar except that: (1) total glycosaminoglycan content was much higher in postnatal animals; (2) chondroitin sulphate was the major glycosaminoglycan after birth; and (3) keratan sulphate, which was barely detectable in the foetal specimens, increased rapidly after birth. Evidence was also obtained for changes in the copolymeric nature of galactosaminoglycans in the inner tissue. These findings, especially the different pattern of age-related changes in outer (presumably non-compressed) and inner (presumably compressed) tissue, suggest that the disc has the capacity to continually modify its composition in response to the mechanical stresses placed on it.