Peripheral axotomy induces only very limited sprouting of coarse myelinated afferents into inner lamina II of rat spinal cord

Abstract

Peripheral axotomy-induced sprouting of thick myelinated afferents (A-fibers) from laminae III-IV into laminae I-II of the spinal cord is a well-established hypothesis for the structural basis of neuropathic pain. However, we show here that the cholera toxin B subunit (CTB), a neuronal tracer used to demonstrate the sprouting of A-fibers in several earlier studies, also labels unmyelinated afferents (C-fibers) in lamina II and thin myelinated afferents in lamina I, when applied after peripheral nerve transection. The lamina II afferents also contained vasoactive intestinal polypeptide and galanin, two neuropeptides mainly expressed in small dorsal root ganglion (DRG) neurons and C-fibers. In an attempt to label large DRG neurons and A-fibers selectively, CTB was applied four days before axotomy (pre-injury-labelling), and sprouting was monitored after axotomy. We found that only a small number of A-fibers sprouted into inner lamina II, a region normally innervated by C-fibers, but not into outer lamina II or lamina I. Such sprouts made synaptic contact with dendrites in inner lamina II. Neuropeptide Y (NPY) was found in these sprouts in inner lamina II, an area very rich in Y1 receptor-positive processes. These results suggest that axotomy-induced sprouting from deeper to superficial layers is much less pronounced than previously assumed, in fact it is only marginal. This limited reorganization involves large NPY immunoreactive DRG neurons sprouting into the Y1 receptor-rich inner lamina II. Even if quantitatively small, it cannot be excluded that this represents a functional circuitry involved in neuropathic pain.

Neuropathy-Induced Spinal GAP-43 Expression Is Not a Main Player in the Onset of Mechanical Pain Hypersensitivity

“…As such, it was thought that low-threshold A fibers directly innervate nociceptionspecific neurons in lamina I, resulting in the gating of nonnoxious stimuli to pain pathways. Although the evidence for A fiber sprouting has been shown to be subject to methodological bias by tracing procedures (Bao et al, 2002;Tong et al, 1999), neuropathy-induced increases in spinal GAP-43 expression still suggest that fiber sprouting occurs within the superficial dorsal horn. Spinal GAP-43 expression was previously reported to be regulated in unmyelinated fibers following nerve injury (Coggeshall et al, 1991).…”

Morphological and phenotypic changes of nociceptive primary afferent neurons following neurotmesis and perineural capsaicin treatment

“…Our findings confirmed previous reports showing the transganglionic labelling of the substantia gelatinosa after the injection of CTB-HRP into injured, but not into intact peripheral nerves , Lekan et al, 1996, Bao et al, 2002. We also demonstrated that neonatal capsaicin treatment produced irreversible loss of small sensory ganglion neurons, and consequently a selective Cfibre deafferentation of the spinal dorsal horn.…”

“…It has also been suggested that this contributes to the development of chronic pain states , 1995, White, 2000. Recently, the sprouting hypothesis has been challenged by showing an increase in the proportion of CTB-horse radish peroxidase (HRP)-labelled small DRG neurons after peripheral nerve transection (Tong et al, 1999 and the co-localization of "injury peptides" such as vasoactive intestinal peptide (VIP) and galanin (GAL), characteristic of injured C-fibre DRG neurons, with choleragenoid in small sensory ganglion neurons and in their central terminations (Bao et al, 2002. Electron microscopic histochemical studies furnished direct evidence for the transport of CTB-HRP in unmyelinated dorsal root axons following peripheral nerve transection .…”

Endophilin A1 regulates dendritic spine morphogenesis and stability through interaction with p140Cap

“…Tissue preparation and immuno-electron microscopy were conducted as described [50]. Briefly, adult mouse was anesthetized with 2% pentobarbital sodium then perfusion-fixed with cold solution containing 0.4% glutaraldehyde (Cat#16220; Electron Microscopy Sciences, Hatfield, PA), 4% PFA (Cat#157-8, Electron Microscopy Sciences) in phosphate buffer (PB; 0.1 M, pH 7.4).…”