Distribution of RET immunoreactivity in the rodent spinal cord and changes after nerve injury

Abstract

RET (for "rearranged during transfection") is a transmembrane tyrosine kinase signaling receptor for members of the glial cell line-derived neurotrophic factor (GDNF) family of ligands. We used RET immunohistochemistry (IHC), double-labeling immunofluorescence (IF), and in situ hybridization (ISH) in adult naïve and nerve-injured rats to study the distribution of RET in the spinal cord. In the dorsal horn, strong RET-immunoreactive (-ir) fibers were abundant in lamina II-inner (II(i)), although this labeling was preferentially observed after an antigen-unmasking procedure. After dorsal rhizotomy, RET-ir fibers in lamina II(i) completely disappeared from the dorsal horn, indicating that they were all primary afferents. After peripheral axotomy, RET-ir in primary afferents decreased in lamina II(i) and appeared to increase slightly in laminae III and IV. RET-ir was also observed in neurons and dendrites throughout the dorsal horn. Some RET-ir neurons in lamina I had the morphological appearance of nociceptive projection neurons, which was confirmed by the finding that 53% of RET-ir neurons in lamina I colocalized with neurokinin-1. GDNF-ir terminals were in close proximity to RET-ir neurons in the superficial dorsal horn. In the ventral horn, RET-ir was strongly expressed by motoneurons, with the strongest staining in small, presumably gamma-motoneurons. Increased RET expression following peripheral axotomy was most pronounced in alpha-motoneurons. The expression and regulation pattern of RET in the spinal cord are in line with its involvement in regenerative processes following nerve injury. The presence of RET in dorsal horn neurons, including nociceptive projection neurons, suggests that RET also has a role in signal transduction at the spinal level. This role may include mediating the effects of GDNF released from nociceptive afferent fibers.

The Neurotrophic Effects of Glial Cell Line-Derived Neurotrophic Factor on Spinal Motoneurons Are Restricted to Fusimotor Subtypes

“…Because Ret expression persists in all MNs into adulthood and appears more highly expressed in small-diameter than large-diameter MNs (Jongen et al, 2007), we tested whether the requirement of GDNF for small-diameter ␥-MNs was specific to development or extended postnatally. We crossed animals expressing a temporally inducible Cre under the ubiquitous ␤-actin promoter (CAGGCre-ER) (Hayashi and McMahon, 2002) to conditional Ret mice, treated with 4-OH-tamoxifen to induce Cre activity every day from P5 to P10 and waited nearly a year to examine fusimotor innervation (Fig.…”

“…Alternatively, early GDNF may be required for the competence of newly differentiated ␥-MNs to respond to intrafusal fiber-derived trophic factors (including GDNF itself). Interestingly, a recent report provided evidence that adult fusimotor MNs express higher levels of Ret than skeletomotor MNs (Jongen et al, 2007). Similarly, developing peroneal MNs, which require GDNF, express higher levels of Ret than GDNF-resistant tibial MNs (Kramer et al, 2006).…”

Autofluorescent Flavoprotein Imaging of Spinal Nociceptive Activity

“…We adjusted the focal plane to the most superficial areas of the rat dorsal spinal cord [i.e., lamina I and II according to Molander et al (1984)]. Since AFI is an optical method and optical responses decrease with increased depth of the focal plane (Sasaki et al, 2002), we hypothesize that the largest contribution of the spinal cord AFI response is generated by projection neurons and local interneurons in these superficial laminae (Jongen et al, 2007;Reinert et al, 2007). However, we cannot exclude that also nerve terminals in the superficial dorsal horn or neurons and nerve terminals in the deeper laminae contribute to the spinal cord AFI response.…”

Changes in Glial Cell Line-derived Neurotrophic Factor Expression in the Rostral and Caudal Stumps of the Transected Adult Rat Spinal Cord

“…7 GDNF levels were significantly different from each other (P \ 0.05), except for the comparison between the 21 dpo and normal group, as well as the 21 and 3 dpo group in the rostral stump. The groups were significantly different from each other (P \ 0.05), except for the comparison between the 3 and 14 dpo group in the caudal stump alpha-motoneurons [38], and topical application of GDNF 30 min after SCI significantly improved motor function and reduced blood-spinal cord barrier (BSCB) breakdown, edema formation, and cell injury at 5 h has been reported [39]. Therefore, GDNF is available to assist in locomotion functional plasticity by exerting behavioral and anatomic neuroprotection following SCI.…”