Neurites from trigeminal ganglion explants grown in vitro are repelled or attracted by tooth-related tissues...

Although neurite attracting factors are present in the developing dental pulp and trigeminal ganglion (TG) axons can respond to such factors, nerve fibres do not enter the tooth pulp until a late developmental stage compared with surrounding tissues supplied by the TG. This suggests that the dental pulp secretes neurite growth inhibitory molecules. Semaphorins represent one group of substances, which can inhibit/repel growing neurites. The aims of the present study were to investigate if dental tissue explants inhibit/repel neurite growth from TGs at some developmental stages in vitro, and if so, to seek evidence for or against a participation of semaphorins in that interaction. By co-culturing mandibular or dental epithelial and mesenchymal tissue explants and TGs in collagen gels, we found that embryonic day 11 (E11) mandibular and E13 dental mesenchymal explants repel neurites from corresponding TGs. Repulsion was replaced by attraction if tissues from late embryonic or early postnatal mice (E17-postnatal day 5) were used. Using semi-quantitative reverse transcription/polymerase chain reaction we showed that a number of semaphorins were expressed by tooth-related mesenchyme collected from embryonic and postnatal mice. The expression of some semaphorins (3A, 3C, 3F, 4F, 5B, 6A, 6B and 6C) was high early in development and then decreased in a temporal pattern that correlated with neurite inhibitory/repulsive effects of dental mesenchyme observed in co-cultures. The expression of other semaphorins increased with development (3B, 4A and 7A), whilst others varied irregularly or remained at a fairly constant level (3E, 4B, 4C, 4D, 4G and 5A). Immunohistochemistry was used to determine if tooth-related nerve fibres possess neuropilins. This revealed that axons surrounding embryonic tooth buds express neuropilin-1, but not neuropilin-2. In postnatal teeth, nerve fibres located within the tooth pulp were immunonegative for neuropilin-1 and neuropilin-2. We conclude that developing mandibular/dental mesenchyme can inhibit/repel neurite growth in vitro. Our results support the hypothesis that semaphorins may be involved in this interaction.