The Sensory and Autonomic Side of Spinal Cord Injury

About Dr. Matt Ramer

Matt Ramer

Matt Ramer

  • 2001-Present-Associate Professor-University of British Columbia and ICORD
  • Post Doc-King’s College London
  • PhD.-Physiology-Queen’s College Kingston, Ontario

Matt Ramer Website

Awards and Funding

Email: ramer@icord.org

Lab Members: A. Gaudet, J. Inskip, A. Scott, L. Soril

Finding Fixes for Injured Nerves

I first became aware of Matt’s research in early 2005. This was catalyzed when he kindly shared excellent IHC images his lab generated using our BDNF and NT-3 antibodies. I was impressed with him and his team’s data and related publications. I did not understand the context of his work and the potential future impact on people suffering peripheral nerve and spinal cord injury (SCI) and wanted to learn more.

He has generously taken the time to open up my view on spinal cord injury (SCI) and what are the challenges in finding therapies and cures. I had equated success soley with restoring mobility. I knew little of the bigger complexites and problems faced by sufferers of SCI.

More than 300,000 people in the United States and Canada suffer from SCI. The ecomonic cost is in the 10s of billions. One of the horrors of SCI is lost mobility.

People with SCI also suffer from a host of problems related to loss of senory and autonomic functions. Sensory and autonomic nerves in the periperal nervous systems (PNS) connect to the spinal cord dorsally. This is different and separate from those controlling movement and motor function. A little know fact is autonomic dysfunctions represent the primary causes of morbidity and mortality following SCI.

So what are the functions that are most important to SCI patients and how should they be prioritized for basic research and drug discovery? Here are two publications that provide insight:

Kim D. Anderson, Ph.D. Targeting Recovery: Priorities of the Spinal Cord-Injured Population. October 1, 2004, 21(10): 1371-1383. doi:10.1089/neu.2004.21.1371.

J A Inskip, L M Ramer, M S Ramer and A V Krassioukov. Autonomic assessment of animals with spinal cord injury: tools, techniques and translation. Spinal Cord advances online publication 10 June 2008; doi: 10.1038/sc.2008.61

The Funding Gap

If we look through the eyes of those suffering from SCI, we know that there are a mryiad of health issues that are outside the problem of lost mobility. I fear the public including those responsible for funding define cure as “the paralyzed can walk”. This is evidenced by a gap between motor vs. sensory/autonomic research and priorities. This gap needs to be closed. The work of Matt and his colleagues represents progress and needs to be supported with funding growth. This backstory highlights how research could feed the discovery of therapies that would answer the recovery priorities of SCI pateints. They, after all, know best.

The Backstory

The story starts in 2000 and highlights Matt’s research at King’s College London. This research was done in collaboration with Dr. Stephen McMahon and Dr. John Priestly.

They showed that regeneration in damaged rat sensory neurons was possible. Injured dorsal roots, treated with nerve growth factor (NGF), neurotrophin-3 (NT3) and glial-cell-line-derived neurotrophic factor (GDNF), but not brain-derived neurotrophic factor (BDNF), resulted in selective regrowth of damaged axons across the dorsal root entry zone and into the spinal cord. Dorsal horn neurons were found to be synaptically driven by peripheral nerve stimulation in rats treated with NGF, NT3 and GDNF, demonstrating functional reconnection. In behavioural studies, rats treated with NGF and GDNF recovered sensitivity to noxious heat and pressure:

Matt S. Ramer, John V. Priestley & Stephen B. McMahon. Functional regeneration of sensory axons into the adult spinal cord. Nature 403, 312-316 (20 January 2000) | doi:10.1038/35002084.

This is a tight rope act. While there is opportunity for regeneration, there are also inhibitors to nerve growth at work. Regeneration becomes more problematic as a function of time. Of the neurotrophins that promote regeneration, NT-3 appears to best at combating the competing inhibitory effects of proteins like NOGO-A. These inhibitory proteins are suspected to be secreted by astrocytes and microglia:

Matt S. Ramer, Ishwari Duraisingam, John V. Priestley, and Stephen B. McMahon. Two-Tiered Inhibition of Axon Regeneration at the Dorsal Root Entry Zone. The Journal of Neuroscience, April 15, 2001, 21(8):2651-2660.

Images: Axon growth 2 weeks after rhizotomy plus immediate NT-3 treatment. A, In intact animals, CTB-labeled terminals are present in lamina I and III, but absent from lamina II. B, Regenerating axons grow along the pial surface of the cord and in the superficial laminae of the gray matter, avoiding the degenerating cuneate fasciculus. C, Dark-field micrograph of B. Scale bar: B, 100 µm. D, Dark-field parasaggital section from a 2 week rhizotomized and NT-3-treated rat. E, Same section as in D, immunostained for CTB. CTB-labeled axons can be seen on the pial surface (arrowheads) and within the cord. Many axons have turned to grow in a rostrocaudal direction but appear to do so in the superficial laminae of the gray matter rather than the white matter. Some individual axons can be traced for up to 2 mm. F, In zones in which the density of regenerated axons is greatest, they form a longitudinal bundle in the gray matter, with few axons in the more superficial white matter (arrows). G, Many axons possess terminal swellings that may be growth cones or termination bulbs. Scale bar: E, 300 µm. The Journal of Neuroscience, April 15, 2001, 21(8):2651-2660.

Through the Looking Glass

Matt and his colleagues continue to gain understanding and refine methods for nerve regeneration. They are also studying plasticity and how these neurons connect to sensory and autonomic neurons in the PNS. This is analogous to re-wiring what was once severed. This would enable restoring of functions important to sufferers of SCI. The related good news is that even partial reconnection enable restoration of these lost functions.

Stepping through the looking glass involves understanding the specific role of these neurons. His recent works include:

Matt Ramer. Anatomical and functional characterization of neuropil in the gracile fasciculus. The Journal of Comparative Neurology. 10.1002/cne.21785.

  • Neurokinin-1 (NK 1) Receptor-Detects a band at 80-90 kDa on Western blots of membranes prepared from cells transfected with the rat substance P receptor (Vigna et al., 1994); stainingin rat spinal cord was blocked by preabsorbing the antiserum with the immunizing peptide (Mantyh et al., 1995)-Dilution 1:2,000
  • Substance P-The distribution of immunoreactivity in rat spinal cord is identical to that described previously (Hunt et al.,1981); in dual-labeling experiments, it labels the same structures as a polyclonal rabbit anti-SP (1:1,000; Peninsula/Bachem; T-4107; data not shown).

Here Matt and his team report on the morphology, inputs, projections, and functional properties of these neurons. Small fusiform and larger lentiform neurons are most abundant in the gracile fasciculus of the cervical and lumbar enlargements and are absent from the cuneate fasciculus and corticospinal tract. Many have dendrites that run along the dorsal pia, and, although in transverse sections these neurons appear isolated from the gray matter, they are also connected to area X by varicose and sometimes loosely fasciculated dendrites. These neurons receive neurochemically diverse, compartmentalized synaptic inputs (primary afferent, intrinsic and descending), half express the substance P receptor, and some project supraspinally. Unlike substantia gelatinosa neurons, they do not express protein kinase C gamma. Functionally, they have small receptive fields, which are somatotopically appropriate with respect to their anterior-posterior position along the neuraxis. They respond to innocuous and/or noxious mechanical stimulation of the distal extremities, and some are prone to central sensitization or windup. Morphologically, neurochemically, and functionally, therefore, these cells most closely resemble neurons in laminae III-VI in the dorsal horn.

Closing Thoughts

There is hope for SCI patients. It is clear that related research and funding needs to expand dramatically beyond the current narrow focus on restored motor function and mobility. The priorities are documented and understood. The story continues. Real progress will be marked by answering these priorities with restored function. Sensing pain, pressure, temperature, etc. where today there is only nothingness. Controlling autonomic functions that pose such a risk to SCI sufferers. I will continue to report the progress of Dr. Matt Ramer and his colleagues. Godspeed to them.

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 reference successful 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 DRGs
was 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, intrathecal
injections of an siRNA against ASIC3 had induced a knockdown of ASIC3 expression in lumbar DRGs.

I will continue to publish updates.

Dr. Matthew Ramer, Neural Regeneration and SCI

Dr. Matthew Ramer

Dr. Matthew Ramer

I dream of the day that people suffering from spinal cord injuries (SCIs) will be restored to full function. The good news is there are researchers and dedicated centers that form the back-bone of SCI repair research.

I am excited and honored to be featuring Dr. Matthew Ramer for this month’s backstory. Matt is a member of ICORD (International Collaboration On Repair Discoveries) at the University of British Columbia. ICORD is an interdisciplinary research centre for the development of effective strategies to promote functional recovery after spinal cord injury. This includes  the discovery and implementation of relevant solutions to improve functional recovery, mobility, community integration and quality of life for people with spinal cord injury.

Matt’s research focus is on the molecular biology of primary sensory nerve cells (neurons). These neurons are responible for sensation. These include touch, pain, temperature, etc. These neurons transmit sensation to the brain via the spinal cord. It is this transmission that enables us to process the sensation. Similar transmissions happen in the case of locomotion.

Matt’s research helps us better understand the mechanism of these transmissions. More importantly, his work includes finding ways to regenerate and repair neurons. These are steps in improving the outcome of sufferes of SCI.

Dicerna Makes Progress

Just wanted to update you on my friends at Dicerna…from in vivo blog…

Wednesday, October 31, 2007

Dicerna Crashes RNAi Party

Life often imitates art, as the saying goes.

In “This Is Spinal Tap,” the classic rock and roll mockumentary chronicling the eponymous band, guitarist Nigel Tufnel famously brags that his amplifiers, unlike conventional ones that max out at a volume of ten, were specially designed to go “one louder.”

“These go to eleven,” he deadpans.

IN VIVO Blog has learned that in the world of RNA interference (RNAi), a new company aims to make some noise of its own not by going louder, but longer, while at the same time circumventing the IP barriers to entry in the exciting field.

Dicerna Pharmaceuticals Inc. is based on technology called Dicer substrate small interfering RNAs developed by co-founders John Rossi, PhD, from the City of Hope National Medical Center’s Beckman Research

Dr. Mark Behlke

Dr. Mark Behlke

Institute and Mark Behlke, MD, PhD, from Integrated DNA Technologies Inc. (IDT). Dicer substrate siRNAs differ from traditional siRNA employed by companies like Alnylam Pharmaceuticals and Merck & Co.’s Sirna Therapeutics in that they are slightly longer oligonucleotides—between 26 and 30 base pairs (bp) versus 21bp for standard siRNA—which then get trimmed down to size once inside the cell. 

Dicerna is expected to announce its $13 million Series A, which will be led by Oxford Bioscience Partners, at some point in November.

Dicerna hopes that its longer molecules not only confer an IP workaround strategy in the hot area of RNA interference therapeutics, but also a pipeline of highly potent drug candidates that will pique the interest of quite a few Big Pharma that have been so far left out of the increasingly expensive but important RNAi arms race.

I will continue to publish updates on this “arms race”.