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Single nerve fibres innervating tooth pulp were isolated from filaments dissected from the inferior alveolar nerve in 17 anaesthetized cats. The fibres were studied to determine whether electrical stimulation of single units produced detectable changes in pulpal blood flow. Single pulpal nerve fibres were electrically stimulated at just above their thresholds and blood flow was recorded with a laser-Doppler flow meter from the pulp of the ipsilateral canine. The thresholds of single fibres in dissected filaments were determined either by recording antidromic action potentials from the tooth or by using a novel technique based on collision. Units that produced blood flow changes were further characterized by recording their response to hot, cold, osmotic and hydrostatic pressure, and mechanical stimulation of exposed dentine and to drying the dentine. Of 93 units isolated, 14 produced changes in pulpal blood flow when stimulated electrically at 1 or 10 Hz. All had conduction velocities (0.8–2.0 m s−1) in the C-fibre range. Ten produced vasodilatation and the remaining four, vasoconstriction. Five of the fibres that produced vasodilatation also responded to the hot stimulus, suggesting that they may form part of an axon reflex or similar mechanism. The four vasoconstrictor units did not respond to any form of stimulus other than electrical and were presumed to be sympathetic post-ganglionic fibres.
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These experiments demonstrate that it is possible to record either an increase or a decrease in pulpal blood flow by electrical stimulation of single pulpal nerve fibres in the cat. Since there was no associated change in arterial blood pressure, it can be assumed that these effects were due to pulpal vasodilatation or vasoconstriction.
The relatively large proportion of the vasodilatation evoked by stimulation of the whole IAN that could be evoked by single fibre stimulation was surprising. This could be explained if, due to the low compliance of the pulpal micro-circulation, there was a maximum vasodilatation that could be produced by the release of neuropeptides following antidromic stimulation and that this maximum effect could be achieved by only a small amount of neuropeptide, not much greater than the quantity released from the terminals of some single units. The volume of the pulp of a lower canine tooth in a young adult cat is approximately 4.2 μl. An alternative explanation is that only a small number of fibres in any one tooth are capable of producing significant vasodilatation. But in that case, we would have to have been very lucky to have found them in the small sample that we isolated. The number of true C-fibres (i.e. those with non-myelinated axons throughout their length, see Cadden et al. 1983) is not known. From the sample we obtained, it would appear that a high proportion of pulpal C-fibres are capable of affecting blood flow in dental pulp. The other possible explanation, that whole nerve stimulation activated both vasodilator and vasoconstrictor fibres and thus produced less vasodilatation than would have occurred with vasodilator stimulation alone, is unlikely since Olgart et al. (1991) found that the vasodilatation produced by electrical stimulation of tooth pulp was not increased following chronic sympathectomy. This is also consistent with the effects we observed with graded stimulation of the inferior alveolar nerve (see Fig. 4).
The observation that some of the units that produced vasodilatation responded to the application of pain-producing stimuli to dentine suggests that they formed part of an axon reflex or axon response mechanism. Consistent with this, Olgart et al. (1991) showed that topical applications of capsaicin or bradykinin to pulp, which are known to excite pulpal C-fibres (Närhi, 1985), produced pulpal vasodilatation in normal, but not in chronically denervated teeth. Heat stimulation did not cause vasodilatation in the present experiments but this can be accounted for by the very short duration of these stimuli. In other experiments (V. Sirimaharaj & B. Matthews, unpublished observations) 30 s heat stimuli have been shown to increase pulpal blood flow. Antidromic stimulation of heat-sensitive pulpal afferents for 30 s in the present experiments also produced vasodilatation.
All of the single fibres that produced a detectable vasodilatation had conduction velocities in the C-fibre range. It was surprising that none of the Aδ fibres caused vasodilatation since, during electrical stimulation of the whole IAN, the threshold for vasodilatation coincided with that of fibres in the Aδ range, as reported previously (Vongsavan & Matthews, 1992a). Also Matthews & Vongsavan (1994) have shown that mechanical stimulation of exposed dentine, which selectively activates A-fibres (Närhi, 1985), evokes pulpal vasodilatation in cats, and many of the nerve terminals in dentinal tubules that are assumed to be sensitive to mechanical stimuli, contain calcitonin gene-related peptide, CGRP (Heyeraas et al. 1993). Olgart et al. (1991) also showed that drilling outer dentine produced vasodilatation in control but not in chronically denervated teeth. Maybe, like the C-fibres, there is only a small number of Aδ fibres that evoke vasodilatation in any one tooth and these were not included in the sample isolated in the current experiments. Alternatively, there may have been overlap in the thresholds of Aδ and C-fibres when the whole IAN was stimulated, so that some C-fibres were recruited at the threshold of vasodilatation. The response of a small number of C-fibres would not be detectable in the compound action potential recorded from a tooth.
Jacobsen & Heyeraas (1997) demonstrated a resting vasodilator tone in ferret dental pulp that was abolished by sectioning the inferior alveolar nerve but not by cervical sympathectomy, indicating that it was of sensory origin. Since pulpal afferents do not discharge impulses spontaneously, and since pulpal blood flow does not change following acute denervation once the acute effects subside (Vongsavan & Matthews, 1992a), it appears that the resting vasodilator tone observed by Jacobsen & Heyeraas may be due to chemical mediators released spontaneously from peripheral sensory nerve terminals.
Lisney and colleagues have made similar observations on vasodilatation produced by Aδ and C-fibre stimulation in rat skin: Jänig & Lisney (1989) showed that electrical stimulation of the peripheral end of the cut saphenous nerve, at intensities that evoked an Aδ wave but not a C-wave in the compound action potential, produced a small transient increase in blood flow. Kolston & Lisney (1993) extended this study by recording blood flow while stimulating filaments that contained single Aδ fibres. They studied 39 fibres and none produced a detectable increase in skin blood flow. To account for these results, they suggested that the amount of neurotransmitter released by a single Aδ fibre might be too small to produce a significant relaxation of local arteriolar smooth muscle, and that simultaneous activation of several such units was necessary to produce a detectable increase in blood flow. Such an explanation could also account for the results of the present experiments.
Lynn et al. (1996) and Gee et al. (1997), who used the method developed for the current experiments to determine the electrical thresholds of single units in nerve filaments, demonstrated that stimulation of single C-fibres could produce detectable vasodilatation in skin. Lynn et al. (1996) showed that, in the pig, antidromic stimulation of single, heat-specific nociceptors produced vasodilatation in the area of the receptive field of the fibre and that stimulation of other classes of C-fibre had no such effect. However, Gee et al. (1997) showed that vasodilatation was also produced by polymodal nociceptors in rats and rabbits. While half the C-fibres that produced vasodilatation in the present experiments were heat sensitive, such fibres have also been shown to respond to chemical stimuli (Närhi et al. 1992).
Of the four vasoconstrictor units identified, none responded to non-electrical stimulation of dentine. This is consistent with them being sympathetic fibres without sensory receptors.
The observation that pulpal blood flow had decreased when recording resumed after exposure and section of the IAN, and in some cases had ceased altogether, indicates that, despite great care to avoid damage to the inferior alveolar artery, the surgery interfered with the arterial supply to the tooth. This observation indicates that blood flow in tissues supplied by the inferior alveolar artery should be recorded routinely before and after exposure of the IAN.