Enamel hypoplasia has been associated with hypoparathyroidism in man and more precisely with the ensuing hypocalcemia. A hypocalcemic state can also be caused by nutritional deficiencies. Most of the experimental models used to assess the role of hypocalcemia on tooth development have employed a diet-induced hypocalcemic state. Rat pups nursed by a dam eating a calcium-deficient diet and weaned on the same diet exhibited a decrease in enamel and dentine deposition and a disturbance of mineralization of these tissues.

After returning to a normal diet, normal mineralized tissues were again formed in the continuously growing incisors. Recently, Ranggård and Norén (1994) reported that diet-induced hypocalcaemia did not greatly affect enamel formation in rat maxillary incisors. However, enamel hypoplasia has been observed in rat incisors after prolonged calcium deficiency.

Parathyroidectomy and thyroparathyroidectomy allows the study of rat incisor development under hypocalcemic and hyperphosphatemic conditions. A previous light- and transmission electron-microscopic study showed that both parathyroidectomy and thyroparathyroidectomy induced enamel defects in the rat incisor. At the beginning of the maturation stage the enamel surface showed marked undulations and numerous surface/subsurface defects. At the electron-microscopic level, granular material was observed in the enamel matrix.

Our aim now was to evaluate by scanning electron microscopy to what extent and in what form the defects observed in early maturation after thyroparathyroidectomy persisted in the mature enamel of the erupted part of the rat incisor. In addition, EDX analyses of iron and calcium could indicate to what extent the development of surface defects affects the deposition of iron pigment in the superficial enamel. Thyroparathyroidectomy, was chosen rather than parathyroidectomy because previous studies had shown that the alterations were always more extensive with thyroparathyroidectomy and that thyroidectomy alone did not induce morphological alterations.

Ten male Wistar rats, 21 days olds (Charles River, France) with a mean body wt of 65 g, were thyroparathyroidectomized under ether anaesthesia. Their weight and ionized blood calcium concentrations were checked 8 and 56 days after surgery. Five sham-operated animals were used as controls. During the experimental period all animals were fed an AO4 diet, not supplemented with carbonate (6 g/kg calcium and 6 g/kg phosphorus; UAR, France) and drank distilled water ad libitum. On day 57, the animals were anaesthetized with 4% chloral hydrate (1 ml/100 g body wt) and fixed by intracardiac perfusion with 1% glutaraldehyde in 0.1 M cacodylate buffer, pH 7.4. The mandibles were dissected out and kept in the same buffer.

The erupted part of the incisor was cut off and glued with cyanoacrylate on its lingual aspect to a small brass cylinder. Specimens were carefully cleaned by brushing lightly with a soft brush under running tap water. After drying overnight they were photographed, sputter-coated with 30-nm gold–palladium, and observed in a Phillips SEM 515 scanning electron microscope operated at 10 kV. After observation of the surface of the enamel, the specimens were cut and ground transversely at their incisal ends, lightly brushed under running tap water, etched twice for 15 sec in 0.1% nitric acid, dried, coated again with gold–palladium and observed in the same microscope operated at 15 kV. The mesiodistal width of the tooth and the mid-labial thickness of enamel and dentine were measured on scanning electron micrographs of the ground and acid-etched transverse plane at the incisal end of the tooth.

EDX analyses of calcium and iron were done with an EDAX® detector/analyser using the Super Quant program and normalizing by element to 100%. Such analyses do not give absolute concentrations. In order to obtain information about the relative amount of iron, it is necessary to compare it with another element. For this purpose we chose calcium, a rather constant component of enamel.

The values of ionized blood calcium concentrations, body wt, incisor length and width, and thickness of enamel and dentine are presented in Table 1. All thyroparathyroidectomized animals had a marked hypocalcemia and reduced body wt, confirming the thyroparathyroidectomized state. The body weight of the thyroparathyroidectomized animals was only 34% of the control value, whereas the width and length of their incisors were reduced to 90% of the control values. The thickness of enamel and dentine was reduced to 83% and 86% of the control values, respectively, indicating that these tissues retained their relative proportions after thyroparathyroidectomy.

Compared with the controls, the incisors of thyroparathyroidectomized animals had a stronger reddish colour, especially concentrated along C-shaped lines, the arms of the C facing incisally (data not shown).

By scanning electron microscopy the surface of normal rat incisor enamel was not completely smooth. The surface roughness showed a distinct pattern in which fine ridges and furrows were orientated longitudinally in the mid-labial region, but deviated towards the cementum–enamel junction in an incisal direction both laterally and mesially. This pattern was also discernible in thyroparathyroidectomized rats, but here it was accentuated by surface defects aligned in the same general direction.

Fig. 2. Scanning electron micrograph of the lateral surface of the erupted part of a control incisor. i, incisal end.

Fig. 3. Scanning electron micrograph of the mid-labial surface of the erupted part of a control incisor. i, incisal end.

Fig. 4. Scanning electron micrograph of the lateral surface of the erupted part of a thyroparathyroidectomized incisor. i, incisal end.

Fig. 5. Scanning electron micrograph of the mid-labial surface of the erupted part of a thyroparathyroidectomized incisor. i, incisal end.

The size and number of enamel surface defects in thyroparathyroidectomized rats varied considerably from one animal to another. Compared to a control, which at this level of magnification presented a rather smooth surface, the enamel of thyroparathyroidectomized rats was more pitted. In most thyroparathyroidectomized animals, larger holes of variable shape were observed; these often contained a plug-like structure.

Fig. 6. Scanning electron micrograph of a higher magnification of the mid-labial surface of the erupted part of a control incisor.

Fig. 7. Scanning electron micrograph of a higher magnification of the mid-labial surface of the erupted part of a different thyro parathyroidectomized incisor.

Fig. 8. Scanning electron micrograph of a higher magnification of the mid-labial surface of the erupted part of a different thyro parathyroidectomized incisor.

Fig. 9. Scanning electron micrograph of a higher magnification of the mid-labial surface of the erupted part of a different thyro parathyroidectomized incisor.

The structure of the mature enamel was observed in transversely sectioned planes close to the incisal end of the icisors. The control incisors showed the typical enamel configuration with uniserial lamellar prism decussation in the inner enamel, incisally inclined parallel prisms in the outer enamel, and an aprismatic superficial layer. The incisors of thyroparathyroidectomized rats exhibited a close to normal enamel structure. The total thickness of the enamel was reduced, but the different layers had retained their relative proportions. The pattern of prism decussation appeared normal, except toward the lateral cementum–enamel junction where the decussation was less distinct or absent.