The Role of Type 1 Metabotropic Glutamate Receptors in the Generation of Dorsal Root Reflexes Induced by Acute Arthritis or the Spinal Infusion of 4-Aminopyridine in the Anesthetized Rat

Abstract

Antidromically propagated action potentials can be recorded in the proximal end of the severed medial articular nerve (MAN) on mechanical stimulation of an inflamed knee in rats and are referred to as dorsal root reflex (DRR) activity. The absence of DRR activity in normal rats suggests that the activity could be the result of hyperexcitability of spinal neurons induced by inflammation. In this study, the role of spinal type 1 metabotropic glutamate (mGlu(1)) receptors in the generation of DRR activity in the MAN during acute knee inflammation was investigated. Four hours after an injection of a mixture of kaolin and carrageenan (k/c) into a knee joint, DRR activity could be evoked in the ipsilateral MAN by mechanical stimulation of the inflamed limb. Spinal application of a selective mGlu(1) receptor antagonist, [RS]-1-Aminoindan-1,5-dicarboxylic acid/UPF 523 (AIDA), or a potent, but less specific mGlu(1) receptor antagonist, LY393053, both depressed the DRR activity significantly. AIDA and LY39053 had no effect on recordings in the MAN from noninflamed control animals. However, spinal administration of AIDA did suppress DRR activity generated by infusion of 4-aminopyridine (4-AP), a K(+) channel blocker, into the dorsal horn of noninflamed animals. These observations suggest that mGlu(1) receptors support the generation of DRR activity in the MAN following sensitization of spinal cord neurons.

Figure 1

(A) Stimulation sites on the lower limb where a 2 N bending force of von Frey fiber was used to evoke DRR activity in animals with an acutely inflamed knee joint. Mechanical stimulation was applied in the order of anterior of knee (a), ankle (b), foot (c) and the lateral part of the knee (d). (B) An example of compound action potentials evoked by von Frey fiber application on the inflamed knee joint and recorded in the proximal end of the cut MAN.

Figure 2

Inhibitory effects of LY393053 on DRR activity in the MAN 4 hours after injection of a mixture of k/c into the knee joint. (A) Control DRR activity recorded from the MAN was evoked in response to a 2 N bending force von Frey fiber applied on knee (a and d), ankle (b) and foot (c), respectively. Examples of the effects of 3 doses of LY393053, 3.6 μmol/L (B), 48.5 μmol/L (C), and 113 μmol/L (D) are shown with DRR activity in the MAN evoked on 4 sites of the lower limb (a to d), respectively (bin width, 1 second). (E) Dose-response curve shown with the averaged percentage of control DRR activity plotted against the concentration shows the dose-dependent inhibitory effect of LY393053 in arthritic rats (P < .001).

Figure 3

Inhibitory effect of AIDA on DRR activity 4 hours after injection of k/c into the knee-joint cavity. (A) Control DRR activity recorded from the MAN induced by a 2 N von Frey fiber applied on knee (a and d), ankle (b) and foot (c) of an arthritic animal. Spinal cord application of 4 μmol/L AIDA (B) for half an hour, there was little effect on the DRR activity. The infusion of 17 μmo/L AIDA (C), or 30 μmol/L (D) into the spinal cord for half an hour inhibited DRR activity recorded from the MAN (bin width, 1 second). (E) Dose response curve for AIDA was constructed using the spike rate of DRR activity (percentage of control) plotted against the calculated concentrations applied to the spinal cord through the microdialysis fiber (P < .001).

Figure 4

Effects of LY393053 (A) and AIDA (B) on the background activity in the MAN in noninflamed control rats. No significant effects on the background activity were observed for these 2 compounds after each dose was infused into spinal cord for 1 hour. Dose response curves were drawn with the spike rate plotted against the drug concentrations. Calculated concentrations applied to the spinal cord through the microdialysis fiber for LY393053 (A) were 0.01, 0.1, 1.85, and 113 μmol/L. The concentrations that were applied for AIDA (B) were 0.03, 0.3, 3, and 30 μmol/L.

Figure 5

The spike rate histograms (left) and the interspike interval histograms (right) constructed from spontaneous antidromic action potentials recorded from the MAN of noninflamed rats (bin width, 1 second). (A) Spike-like background activity was recorded from the MAN during microdialysis infusion of aCSF into the spinal cord for about 1 hour. The irregular, antidromic, multi-unit activity had a mean firing rate of 8/sec. Interspike interval histogram (A′) showed the number of counts at each interval with the peak interval at 4 milliseconds. (B) Spinal microdialysis administration of 4-AP (at the concentration of about 27.7 μmol/L for 1 hour) the frequency of spike activity was greatly increased with a mean firing rate of 26/sec. (B′) The peak interspike interval at 4 milliseconds increased in magnitude to about 5 times that of the aCSF control (A′). (C) and (C′) Microdialysis coinfusion of 4-AP and AIDA (17 μmol/L) for about 1 hour reversed the effect of 4-AP. The mean firing rate decreased to the level of the aCSF control (8.29/sec). The interspike interval histogram (C′) returned to the control level.

Figure 6

(A) Dose response curve of the firing rate in the MAN plotted against 4-AP concentrations showing that 4-AP increases the frequency of the spontaneous DRR activity in a dose-dependent manner. (B) A bar chart showing the effects of 4-AP (middle), and the coapplication of 4-AP plus AIDA (right), on the mean firing rate of spontaneous background activity recorded from the MAN. Spinal cord application of 4-AP (for 1 hour) at the concentration of 27.7 μmol/L increased the mean spike rate in DRR recorded from the MAN significantly (P ≤.05). Coinfusion of 4-AP with AIDA (at the concentration of 17 μmol/L) for about 1 hour reversed the increased spike frequency induced by 4-AP back to the control level. Data used in A and B were from different groups of animals. Accumulating doses of 4-AP were applied to the spinal cord in (A), whereas in (B) only 1 dose of 4-AP at 27.7 μmol/L was applied for 1 hour, which was then followed by the coapplication of 4-AP and AIDA.