Functional downregulation of P2X3 receptor subunit in rat sensory neurons reveals a significant role in chronic neuropathic and inflammatory pain.
The excitation of nociceptive sensory neurons by ATP released in injured tissue is believed to be mediated partly by P2X3 receptors. Although an analysis of P2X3 knock-out mice has revealed some deficits in nociceptive signaling, detailed analysis of the role of these receptors is hampered by the lack of potent specific pharmacological tools. Here we have used antisense oligonucleotides (ASOs) to downregulate P2X3 receptors to examine their role in models of chronic pain in the rat. ASOs and control missense oligonucleotides (180 microg/d) were administered intrathecally to naive rats for up to 7 d via a lumbar indwelling cannula attached to an osmotic minipump. Functional downregulation of the receptors was confirmed by alphabeta-methylene ATP injection into the hindpaw, which evoked significantly less mechanical hyperalgesia as early as 2 d after treatment with ASOs relative to controls. At this time point, P2X3 protein levels were significantly downregulated in lumbar L4 and L5 dorsal root ganglia. After 7 d of ASO treatment, P2X3 protein levels were reduced in the primary afferent terminals in the lumbar dorsal horn of the spinal cord. In models of neuropathic (partial sciatic ligation) and inflammatory (complete Freund's adjuvant) pain, inhibition of the development of mechanical hyperalgesia as well as significant reversal of established hyperalgesia were observed within 2 d of ASO treatment. The time course of the reversal of hyperalgesia is consistent with downregulation of P2X3 receptor protein and function. This study demonstrates the utility of ASO approaches for validating gene targets in in vivo pain models and provides evidence for a role of P2X3 receptors in the pathophysiology of chronic pain.
Barclay
Jane
J
Novartis Institute for Medical Sciences, London WC1E 6BS, United Kingdom.
Patel
Sadhana
S
Dorn
Gabriele
G
Wotherspoon
Glen
G
Moffatt
Sarah
S
Eunson
Louise
L
Abdel'al
Samir
S
Natt
Francois
F
Hall
Jonathan
J
Winter
Janet
J
Bevan
Stuart
S
Wishart
William
W
Fox
Alyson
A
Ganju
Pam
P
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives; D000818:Animals; D004195:Disease Models, Animal; D015536:Down-Regulation / Q000502:physiology; D005620:Freund's Adjuvant; D006614:Hindlimb; D006930:Hyperalgesia / Q000139:chemically induced / Q000188:drug therapy / Q000503:physiopathology*; D007249:Inflammation / Q000139:chemically induced / Q000503:physiopathology*; D007278:Injections, Spinal; D008026:Ligation; D008297:Male; D009475:Neurons, Afferent / Q000166:cytology / Q000378:metabolism*; D016376:Oligonucleotides, Antisense / Q000008:administration & dosage / Q000494:pharmacology; D010147:Pain Measurement; D021122:Protein Subunits; D012333:RNA, Messenger / Q000037:antagonists & inhibitors / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D017208:Rats, Wistar; D018048:Receptors, Purinergic P2 / Q000235:genetics / Q000378:metabolism*; D058477:Receptors, Purinergic P2X3; D020426:Sciatic Neuropathy / Q000188:drug therapy / Q000503:physiopathology*
D016376:Oligonucleotides, Antisense; C546409:P2rx3 protein, mouse; C546410:P2rx3 protein, rat; D021122:Protein Subunits; D012333:RNA, Messenger; D018048:Receptors, Purinergic P2; D058477:Receptors, Purinergic P2X3; D000255:Adenosine Triphosphate; D005620:Freund's Adjuvant; C002630:alpha,beta-methyleneadenosine 5'-triphosphate