Reduced inflammatory hyperalgesia with preservation of acute thermal nociception in mice lacking cGMP-dependent protein kinase I

Abstract

cGMP-dependent protein kinase I (PKG-I) has been suggested to contribute to the facilitation of nociceptive transmission in the spinal cord presumably by acting as a downstream target of nitric oxide. However, PKG-I activators caused conflicting effects on nociceptive behavior. In the present study we used PKG-I ؊/؊ mice to further assess the role of PKG-I in nociception. PKG-I deficiency was associated with reduced nociceptive behavior in the formalin assay and zymosan-induced paw inflammation. However, acute thermal nociception in the hot-plate test was unaltered. After spinal delivery of the PKG inhibitor, Rp-8-Br-cGMPS, nociceptive behavior of PKG-I ؉/؉ mice was indistinguishable from that of PKG-I ؊/؊ mice. On the other hand, the PKG activator, 8-Br-cGMP (250 nmol intrathecally) caused mechanical allodynia only in PKG-I ؉/؉ mice, indicating that the presence of PKG-I was essential for this effect. Immunofluorescence studies of the spinal cord revealed additional morphological differences. In the dorsal horn of 3-to 4-week-old PKG-I ؊/؊ mice laminae I-III were smaller and contained fewer neurons than controls. Furthermore, the density of substance P-positive neurons and fibers was significantly reduced. The paucity of substance P in laminae I-III may contribute to the reduction of nociception in PKG-I ؊/؊ mice and suggests a role of PKG-I in substance P synthesis.spinal cord ͉ substance P ͉ nitric oxide ͉ pain T he second messenger cGMP is formed by activation of soluble and particulate guanylyl cyclases and has several targets, including cGMP-dependent protein kinase I (PKG-I) and PKG-II, of which PKG-I is expressed in the spinal cord (1, 2). Spinally delivered PKG inhibitors reduce formalin-induced nociceptive behavior in rats (3, 4), suggesting that PKG-I plays an important role in spinal nociceptive processing. It has been speculated that PKG-I mediates hyperalgesic effects of nitric oxide (NO) (5). This idea is supported by the observation that PKG-I inhibition causes a reduction of thermal hyperalgesia induced by injection of the NO donor, NOC-12 (6). Endogenous NO is produced by NO synthases, of which neuronal nitric oxide synthase (nNOS) is activated and up-regulated after N-methyl-D-aspartate receptor stimulation (7-10). NO probably acts as a retrograde messenger (11, 12) at nociceptive synapses, i.e., it is released from the postsynaptic neuron, diffuses back to the presynaptic neuron, and stimulates guanylyl cyclases. The latter step links NO to cGMP production and PKG-I activation. Because inhibition of NOS activity reduces nociception (13,14), the release of NO is thought to contribute to the development of hyperexcitability of nociceptive neurons under certain circumstances. Under the premise that PKG-I is a mediator of NO at nociceptive synapses, one would expect that PKG-I activation also causes hyperalgesia. However, effects of the spinally delivered PKG activator 8-Br-cGMP have been conflicting (15,16). (See Supporting Text, which is published as supporting information on the PNAS web site....

Cysteine-Rich Protein 2, a Novel Downstream Effector of cGMP/cGMP-Dependent Protein Kinase I-Mediated Persistent Inflammatory Pain

“…The expression of cGKI in the spinal cord is upregulated after noxious stimulation, and intrathecal administration of cGKI inhibitors provided profound antinociceptive effects in various animal models of pain Schmidtko et al, 2003;Song et al, 2006). The essential contribution of cGKI to the mechanisms underlying pain hypersensitivity is further supported by strongly reduced inflammatory hyperalgesia in cGKI knock-out mice (Tegeder et al, 2004).…”

“…After baseline measurements, the cGMP analogs 8-Br-cGMP or 8-pCPT-cGMP (Biolog), dissolved in artificial CSF (ACSF; 141.7 mM Na ϩ , 2.6 mM K ϩ , 0.9 mM Mg 2ϩ , 1.3 mM Ca 2ϩ , 122.7 mM Cl Ϫ , 21.0 mM HCO 3 Ϫ , 2.5 mM HPO 4 2Ϫ , 3.5 mM dextrose, bubbled with 5% CO 2 in 95% O 2 to adjust pH to 7.2), were intrathecally injected in a volume of 2.5 l through the catheter followed by 2.5 l of ACSF. Based on previous results in mice (Tegeder et al, 2004) and rats (Tegeder et al, 2002), 250 and 25 nmol of 8-BrcGMP were considered as high and low dose, respectively. The high (25 nmol) and low (1.25 nmol) dose of 8-pCPT-cGMP, which has a higher membrane permeability and phosphodiesterase stability than 8-Br-cGMP (Butt et al, 1992;Schwede et al, 2000) was determined in preliminary experiments.…”

Sphingosine 1-Phosphate Modulates Spinal Nociceptive Processing

“…The formalin assay (18), and the hot plate test (19) were performed as described previously. The observer was unaware of the treatments in all tests.…”

CNGA3: A Target of Spinal Nitric Oxide/cGMP Signaling and Modulator of Inflammatory Pain Hypersensitivity

“…In fact, several studies revealed that NO donors and cGMP analogs, in addition to their well characterized pronociceptive effects, may also inhibit pain (Iwamoto and Marion, 1994;Pehl and Schmid, 1997;Tegeder et al, 2002Tegeder et al, , 2004Schmidtko et al, 2008b). Interestingly, the antinociceptive effects of cGMP analogs were not antagonized by coadministration with a cGKI inhibitor and were also observed in cGKI Ϫ/Ϫ mice, suggesting that cGKI-independent mechanisms are involved.…”

“…The contribution of cGKI to pain sensitization is reflected by the reduced inflammatory hyperalgesia in cGKI Ϫ/Ϫ mice (Tegeder et al, 2004), and by antinociceptive effects of cGKI inhibitors (Tao et al, 2000;Schmidtko et al, 2003). However, there is accumulating evidence that cGMP may also exert antinociceptive effects within the spinal cord independently from cGKI activation (Iwamoto and Marion, 1994;Tegeder et al, 2002Tegeder et al, , 2004Schmidtko et al, 2008b).…”