Some lumbar sympathetic neurons develop a glutamatergic phenotype after peripheral axotomy with a note on VGLUT2-positive perineuronal baskets

Glutamate is the main excitatory neurotransmitter in the nervous system, including in primary afferent neurons. However, to date a glutamatergic phenotype of autonomic neurons has not been described. Therefore, we explored the expression of vesicular glutamate transporter (VGLUT) types 1, 2 and 3 in lumbar sympathetic chain (LSC) and major pelvic ganglion (MPG) of naïve BALB/C mice, as well as after pelvic nerve axotomy (PNA), using immunohistochemistry and in situ hybridization. Colocalization with activating transcription factor-3 (ATF-3), tyrosine hydroxylase (TH), vesicular acetylcholine transporter (VAChT) and calcitonin gene-related peptide was also examined. Sham-PNA, sciatic nerve axotomy (SNA) or naïve mice were included. In naïve mice, VGLUT2-like immunoreactivity (LI) was only detected in fibers and varicosities in LSC and MPG; no ATF-3-immunoreactive (IR) neurons were visible. In contrast, PNA induced upregulation of VGLUT2 protein and transcript, as well as of ATF-3-LI in subpopulations of LSC neurons. Interestingly, VGLUT2-IR LSC neurons coexpressed ATF-3, and often lacked the noradrenergic marker TH. SNA only increased VGLUT2 protein and transcript in scattered LSC neurons. Neither PNA nor SNA upregulated VGLUT2 in MPG neurons. We also found perineuronal baskets immunoreactive either for VGLUT2 or the acetylcholinergic marker VAChT in non-PNA MPGs, usually around TH-IR neurons. VGLUT1-LI was restricted to some varicosities in MPGs, was absent in LSCs, and remained largely unaffected by PNA or SNA. This was confirmed by the lack of expression of VGLUT1 or VGLUT3 mRNAs in LSCs, even after PNA or SNA. Taken together, axotomy of visceral and non-visceral nerves results in a glutamatergic phenotype of some LSC neurons. In addition, we show previously non-described MPG perineuronal glutamatergic baskets.Fil: Brumovsky, Pablo Rodolfo. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Austral; Argentina. Univeristy of Pittsburgh. School of Medicine; Estados UnidosFil: Seroogy, Kim B.. University of Cincinnati; Estados UnidosFil: Lundgren, Kerstin H.. University of Cincinnati; Estados UnidosFil: Watanabe, Masahiko. Hokkaido University School of Medicine; JapónFil: Hökfelt, Tomas. Karolinska Huddinge Hospital. Karolinska Institutet; SueciaFil: Gebhart, G.F.. Univeristy of Pittsburgh. School of Medicine; Estados Unido

Axotomy of tributaries of the pelvic and pudendal nerves induces changes in the neurochemistry of mouse dorsal root ganglion neurons and the spinal cord

Using immunohistochemical techniques, we characterized changes in the expression of several neurochemical markers in lumbar 4-sacral 2 (L4–S2) dorsal root ganglion (DRG) neuron profiles (NPs) and the spinal cord of BALB/c mice after axotomy of the L6 and S1 spinal nerves, major tributaries of the pelvic (targeting pelvic visceral organs) and pudendal (targeting perineum and genitalia) nerves. Sham animals were included. Expression of cyclic AMP-dependent transcription factor 3 (ATF3), calcitonin gene-related peptide (CGRP), transient receptor potential cation channel subfamily V, member 1 (TRPV1), tyrosine hydroxylase (TH) and vesicular glutamate transporters (VGLUT) types 1 and -2 was analysed seven days after injury. L6–S1 axotomy induced dramatic de novo expression of ATF3 in many L6–S1 DRG NPs, and parallel significant downregulations in the percentage of CGRP-, TRPV1-, TH- and VGLUT2-immunoreactive (IR) DRG NPs, as compared to their expression in uninjured DRGs (contralateral L6–S1-AXO; sham mice); VGLUT1 expression remained unaltered. Sham L6–S1 DRGs only showed a small ipsilateral increase in ATF3-IR NPs (other markers were unchanged). L6–S1-AXO induced de novo expression of ATF3 in several lumbosacral spinal cord motoneurons and parasympathetic preganglionic neurons; in sham mice the effect was limited to a few motoneurons. Finally, a moderate decrease in CGRP- and TRPV1-like-immunoreactivities was observed in the ipsilateral superficial dorsal horn neuropil. In conclusion, injury of a mixed visceral/non-visceral nerve leads to considerable neurochemical alterations in DRGs matched, to some extent, in the spinal cord. Changes in these and potentially other nociception-related molecules could contribute to pain due to injury of nerves in the abdominopelvic cavity.Fil: Mccarthy, Carly Jane. Universidad Austral. Facultad de Ciencias Biomédicas; ArgentinaFil: Tomasella, María Eugenia. Universidad Austral. Facultad de Ciencias Biomédicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Malet, Mariana. Universidad Austral. Facultad de Ciencias Biomédicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Seroogy, Kim B.. University of Cincinnati; Estados UnidosFil: Hökfelt, Tomas. Karolinska Huddinge Hospital. Karolinska Institutet; SueciaFil: Villar, Marcelo Jose. Universidad Austral. Facultad de Ciencias Biomédicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Gebhart, G.F.. University of Pittsburgh; Estados UnidosFil: Brumovsky, Pablo Rodolfo. University of Pittsburgh; Estados Unidos. Universidad Austral. Facultad de Ciencias Biomédicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Localization and Changes in NADPH-Diaphorase Reactivity and Nitric Oxide Synthase Immunoreactivity in Rat Pulp Following Tooth Preparation

Inflammatory changes in the dental pulp are accompanied by release of a wide variety of chemical mediators. Nitric oxide, an oxidative free radical produced by the enzyme nitric oxide synthase (NOS), has been implicated in multiple inflammatory processes, which makes it a suitable marker for changes which likely occur following tooth pulp insult. Since limited information on nitric oxide in the pulp is available, it is necessary first to examine relative distributions of NOS in uninflamed and inflamed rat pulp. We accomplished this by characterizing regions of nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) activity and the distribution of both macrophage NOS (macNOS) and neuronal NOS (nNOS) immunoreactivity in normal and inflamed rat molar pulp at multiple time points. The results showed that: (1) deep cavity preparation on the mesial surface of the molar produced a time-dependent inflammation, with acute inflammation early progressing to chronic, granulomatous inflammation with necrosis later that spread preferentially down the mesial root; (2) control (non-prepared) teeth showed a relatively faint and homogeneous distribution of NADPH-d and macNOS reactivity but no discernible nNOS reactivity; (3) inflamed teeth displayed localized increased intensity of NADPH-d and macNOS reactivity surrounding the inflamed area of pulp, but no increased nNOS activity; (4) pulp vessels supplying the inflamed area showed increased NADPH-d reactivity, but no increased macNOS or nNOS reactivity; and (5) neither NADPH-d, macNOS, nor nNOS reactivity was observed in pulpal nerves. Therefore, nitric oxide may mediate the pulpal inflammatory response through its effects on the paralesional pulp tissue and surrounding endothelial/vascular structures.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/67744/2/10.1177_00220345990780100301.pd