Cannabinoid modulation of nociception and nociceptor activity during inflammation.

University of Minnesota Ph.D. dissertation. June 2009. Major: Neuroscience. Advisor: Donald Anthony Simone. 1 computer file (PDF); x, 235 pages.Previous studies have demonstrated that peripherally-administered cannabinoids at the site of injury produce antinociception in animal models of acute and persistent pain. Peripheral cannabinoid one (CB1) receptor-mediated antinociception has been attributed to CB1 receptors located on nociceptive DRG neurons and their peripheral nerve terminals. Although these studies suggest that activation of peripheral CB1 receptors located on nociceptive nerve terminals produces antinociception, how cannabinoids modulate nociceptor activity is not known. The overall aim of this thesis was to relate the behavioral antinociceptive effects of locally-administered cannabinoids with changes in the response properties of nociceptors during non-inflamed and inflamed conditions. It was hypothesized that activation of peripheral CB1 receptors attenuated nociception and nociceptor activity only during inflammation. In behavioral studies, intraplantar administration of complete Freund's adjuvant (CFA), but not saline, produced mechanical allodynia, mechanical hyperalgesia, and heat hyperalgesia. Activation of peripheral CB1 receptors produced antiallodynia and antihyperalgesia following inflammation, but did not alter nociception during non-inflamed conditions. In electrophysiological studies, only cutaneous nociceptors (Adelta and C) from inflamed skin were sensitized, and not Abeta mechanoreceptors. Local administration of CB1 receptor agonists attenuated mechanically-evoked responses of Adelta nociceptors from inflamed skin, but did not alter the evoked responses of Adelta nociceptors from non-inflamed skin. The responses of C nociceptors and Abeta mechanoreceptors from either non-inflamed or inflamed skin were not altered following local administration of cannabinoids. Our results demonstrated that peripherally-mediated cannabinoid antinociception through CB1 receptors is mediated, at least in part, by attenuation of Adelta nociceptor activity. The results from the present studies suggest that peripherally-acting CB1 receptor agonists could be administered alone or co-administered with other analgesic drugs to treat acute and persistent pain in humans and animals

Parainfluenza 3-Induced Cough Hypersensitivity in the Guinea Pig Airways

<div><p>The effect of respiratory tract viral infection on evoked cough in guinea pigs was evaluated. Guinea pigs were inoculated intranasally with either parainfluenza type 3 (PIV3) and cough was quantified in conscious animals. The guinea pigs infected with PIV3 (day 4) coughed nearly three times more than those treated with the viral growth medium in response to capsaicin, citric acid, and bradykinin. Since capsaicin, citric acid, and bradykinin evoked coughing in guinea pigs can be inhibited by drugs that antagonize the transient receptor potential cation channel, subfamily V, member 1 (TRPV1), it was reasoned that the virally-induced hypertussive state may involve alterations in TPRV1 activity. PIV3 infection caused a phenotypic switch in tracheal nodose Aδ “cough receptors” such that nearly 50% of neurons began to express, de novo, TRPV1 mRNA. There was also an increase TRPV1 expression in jugular C-fiber neurons as determined by qPCR. It has previously been reported that tracheal-specific nodose neurons express the BDNF receptor TrkB and jugular neurons express the NGF receptor TrkA. Jugular neurons also express the artemin receptor GFRα3. All these neurotrophic factors have been associated with increases in TRPV1 expression. In an ex vivo perfused guinea pig tracheal preparation, we demonstrated that within 8 h of PIV3 infusion there was no change in NGF mRNA expression, but there was nearly a 10-fold increase in BDNF mRNA in the tissue, and a small but significant elevation in the expression of artemin mRNA. In summary, PIV3 infection leads to elevations in TRPV1 expression in the two key cough evoking nerve subtypes in the guinea pig trachea, and this is associated with a hypertussive state with respect to various TRPV1 activating stimuli.</p></div

Sequence of primers used for analysis of guinea pig ganglia receptor transcripts.

<p>Sequence of primers used for analysis of guinea pig ganglia receptor transcripts.</p

Guinea pig cough responses to inhaled capsaicin (0.1, 1, 3, 10 μM), citiric acid (CA; 0.01, 0.1, 0.3 M) and bradykinin (BK; 0.1 mg/ml) 4 d after viral inoculation.

<p>Cough was confirmed in conscious guinea pigs based on changes in pressure and visual confirmation. Comparison of the average number of coughs between infected animals and controls were compared. A) Capsaicin: Control, n = 4; PIV3: n = 4. B) CA: Control, n = 8; PIV3, n = 8. D) Bradykinin: Control, n = 10; PIV3, n = 10. *Significantly different from control (p<0.05).</p

TRPV1 expression in the jugular ganglia in control animals compared to those inoculated with PIV3 (day 4).

<p>Increased expression of TRPV1 (qPCR) following PIV3 infection for 4 d. Control: n = 7, 14 ganglia; PIV3: n = 7, 14 ganglia. *Significantly different from control (p<0.05).</p

PIV3 infection of epithelium in guinea pig trachea 4 d after inoculation with virus or vehicle (control).

<p>A) PIV3 staining of tracheal epithelium from a vehicle-treated control animal. B) PIV3 staining of tracheal epithelium following viral infection in vivo. Bar = 100 μm. C) Each lane represents RT-PCR amplification with primers for PIV3 genes: nucleocapsid protein (NP), fusion protein (FP), hemagglutinin-neuraminidase (HN), and a positive control- guinea pig β-actin (β).</p

Sequence of primers used for PCR analysis of PIV3.

<p>Sequence of primers used for PCR analysis of PIV3.</p

Sequence of primers used for qPCR analysis of guinea pig neurotrophin transcripts.

<p>Sequence of primers used for qPCR analysis of guinea pig neurotrophin transcripts.</p

Percentage of guinea pig tracheal-specific nodose neurons that express TRPV1 in controls compared to those inoculated with PIV3 (day 4).

<p>Top, representative mRNA expression of TRPV1 from neurons. Bottom, total TRPV1 mRNA expression in infected animals (49.48 ±5.1%) compared to controls (16.28 ±4.0%). Control: n = 11, 86 neurons; PIV3: n = 10, 97 neurons. *Significantly different from control (p<0.05).</p