Abstract Aim To quantify the expression of calcitonin gene‐related peptide (CGRP), substance P (SP), neurokinin A (NKA), neuropeptide Y (NPY) and vasoactive intestinal polypeptide (VIP) in healthy and inflamed human dental pulp tissue. Methodology Six pulp samples were obtained from teeth having a clinical diagnosis of acute irreversible pulpitis. Another 12 pulp samples were obtained from premolars where extraction was indicated for orthodontic purposes. In six of these premolar teeth inflammation was induced by mechanical pulp exposure prior to sample collection. All samples were processed and 125 I‐labelled; neuropeptides were quantified by competition assays. anova and Mann–Whitney's ( post hoc ) tests were used to establish statistically significant differences between the groups. Results Expression of five neuropeptides was found in all human pulp samples. Statistical analysis revealed a significantly higher ( P < 0.05) expression of CGRP, SP, NKA and NPY in both inflammatory conditions compared with healthy pulp control values. VIP expression remained stable during the inflammatory conditions. Conclusion Expression of CGRP, SP and NKA released from C‐fibres and NPY released from sympathetic fibres is significantly higher in the inflamed human pulp compared with healthy pulp. Expression of VIP released from parasympathetic fibres is not increased during the inflammatory conditions of human dental pulp.