ACIC3 Receptors Knockdown in vivo

Researchers using siRNA complexed with our i-Fect ™ transfection regent have successfully knocked down ASIC3 Receptors in vivo. This publication joins the growing parade (starting with Luo et al, 2005) that refererence successuful modulation of receptors involved in pain using siRNA complexes.

These studies all share animal behavior studies showing a marked change in response to pain stimuli after treatment.

In this study, Dr. Eric Lingueglia and his team found Peripheral ASIC3 channels are thus essential sensors of acidic pain and integrators of molecular signals produced during inflammation where they contribute to primary hyperalgesia.

Emmanuel Deval, Jacques Noël, Nadège Lay, Abdelkrim Alloui, Sylvie Diochot, Valérie Friend, Martine Jodar, Michel Lazdunski and Eric Lingueglia. ASIC3, a sensor of acidic and primary inflammatory pain. The EMBO Journal advance online publication 16 October 2008; doi: 10.1038/emboj.2008.213

Cy3-labelled siRNA no. 1121 and its corresponding scramble (no. 1121S; GCTCACACTACGCAGAGAT) synthesized by MWG Biotech (Germany) were injected in rats by intrathecal bolus to the lumbar region of the spinal cord once a day for 3 days before the induction of inflammation with CFA. Each 10-ml injection corresponded to 2 mg of siRNA complexed with i-Fect siRNA transfection reagent (Neuromics) at a ratio of 1:4 (w:v) (Luo et al, 2005), following the supplier’s suggested protocol. siRNA uptake in lumbar DRGswas monitored by fluorescence microscopy on cryostat sections 24 h after a single intrathecal injection.

Here’s a synopsis of results:
Inflammation was produced by CFA injection, which led to primary heat hyperalgesia, and this hyperalgesia was drastically reduced by the ASIC3 blocker APETx2 injected subcutaneously, which only access cutaneous nociceptors. It was also drastically reduced when, before triggering the inflammation state, intrathecalinjections of an siRNA against ASIC3 had induced a knockdown of ASIC3 expression in lumbar DRGs.

siRNA-mediated gene silencing

Dr. Josephine Lai (Professor of Pharmacology, University of Arizona) is a pioneer in developing experimental designs and methods for delivering siRNA to the CNS for gene expression analysis.She and her team have documented these in the publication:


Modulating Sensory Systems Using RNAi(pdf - 187Kb)© 2007 Lai


For researchers desiring to effectively deliver siRNA to the CNS for gene expression analysis of specific receptors, this publication offers proven methods. These include:

• The Choice of siRNA


• Choosing and Optimizing Transfection Reagents for siRNA Delivery to the Nervous System
  
• Delivery Systems-Microinjection and Infusion (using mini-osmotic pumps)
  
• Validation

We will continue to track advances by Dr. Lai and team.