Nerve injury & Motoneurons

1. Nerve injury &Motoneurons Core concepts: Loss of trophic support Excitotoxicity

2. Causes of motoneuron death NOCD Early postnatal injury Disease SMA ALS PMN

3. ~5000 affected in UK 1-2/100,000 new cases p.a. Affects men > women Onset >40 years of age 55-65 yrs old most commonly affected Genetic Not possible to prevent onset Symptoms Progressive muscle weakness & wasting Hands, arms & legs usually affected first May get spasticity, painful cramps & loss of balance Affects vocal & respiratory muscles Motoneuron disease

4. Clinical syndromes: Spinal Muscular Atrophy • Hereditary condition defect in SMN gene (Chromosome 5q12.2-q13) • Insufficient peripheral motor sprouting • Increased motoneuron activity – excitotoxicity?

5. Clinical syndromes:Age related motoneuron loss Senile Muscular Atrophy • Affects 15% of elderly population • 10-20% motoneuron loss • Motoneurons show signs of damage •  CGRP, GAP43, p75 •  trkB, trkC;

6. ALS • Affects UMN/LMNs • Mechanisms • 10% Hereditary (SOD1) • Oxidative stress • EAA toxicity • Glial EAAT2 abnormal •  Glutamate in CNS •  Glutamate in CSF • NOS, Cox 2 induced • Defect in RA pathway

7. Goals of neuromuscular disease research • Prevent death • Maintain phenotype • Repair neuronal damage

8. Trophic factors and motoneuron survival BDNF, NT3, NT4 GDNF, NTN, PSN LIF, CNTF, CT1 FGFs RA HGF IGF1 BDNF, NT3, NT4 GDNF LIF, CNTF FGF5, bFGF IGF1, IGF2

9. NOCD & trophic factor knockout NGF- trk A No motoneuron loss BDNF/NT4 - trk B No motoneuron loss NT3 - trk C No motoneuron loss p75 No motoneuron loss BDNF-NT4 double KO No motoneuron loss trk B/C double KO No motoneuron loss CNTF No motoneuron loss CNTFR 40-50% LOSS LIFR LOSS gp130 40% LOSS GDNF- GFR1 20-40% LOSS GFR2 No motoneuron loss c-RET Significant loss

10. Motoneuron survival depends on age P0 90% P3 80% P4 30% P5 0% and post-operative survival time

11. Neonatal motoneuron death depends on lesion site • Motoneuron loss also depends on duration of deprivation • P0 axotomy: 1% survival • P0 crush: 10-30% survival • Delayed reinnervation P5 & P10 crush 60% survival YES   YES Yes

12. Is motoneuron death due to axon damage per se or target deprivation? • Motoneuron loss is regulated by target deprivation • can be induced by NMJ blockade at birth (maintains MN immaturity) injury induced release of glutamate (kills immature MNs) Can be mimicked by NMDA injection

13. Motoneuron death is regulated by target contact Growing mode Transmitting mode Muscle induced neuro-transmitter release Ach

14. Motoneuron death is regulated by target contact Growing mode Preserved immature state Death by glutamate excito- toxicity Prevention of neuromuscular interaction No induced neuro-transmitter release

15. Motoneuron maturation Growing neurone transmitting neurone

16. Biochemical  expression, Reg2, HSP27, GFR1, p75, CGRP, CB, gp130, trkC, CNTFR  expression GAP43, c-Jun, NOS, NR1, NR2B, GAL, mRNAs for LIF, trkB, c-RET Physiological  neuronal activity abnormal reflex patterns  dendrite number, altered morphology Neonatal axotomy: effects on surviving neurons

17. Neurotrophic support Preventing excitotoxicity Important Outcomes Permanent survival Rescued motoneurons must reinnervate muscles Muscles must develop adequate force on reinnervation Spinal circuits must be re-established Pharmacological manipulations that rescue dying motoneurons

18. Neurotrophic support 1 week 2 weeks combination nerve + s.c P3 sciatic cut + single dose NTF treatment to injury site

19. Transient rescue BDNF < 3 weeks NT3 < 2 weeks NT4 < 1week CNTF < 2 weeks LIF < 2 weeks GDNF combinations Restore ChAT levels Permanent rescue GDNF (AAV) Deleterious NGF (activity dependent) High dose BDNF receptor desensitisation or activity dependent Neonatal neurotrophic support

20. Adult motoneuron death YES   NO/ YES -delayed NO

21. Loss of normal function Loss of reflex function Soma atrophy Motor c.v.   ChAT  transmitter receptors No cell death* VRA Repeated nerve injury Regeneration  GAP43, c-Jun  CGRP, GAL,  REG2  HSP27  c-Ret, GFR1, LIFR, p75, CNTFR, trkB  gp130,  trkC Adult nerve injury

22. Exogenous NTFs reverse effects of injury & aid regeneration NGF, BDNF, GDNF, NT4 * **  trkB GFR1 p75 Regeneration GAP43, CGRP Tubulin CNTF,  NT3 p75  apoptosis in Schwann cells   regeneration

23. Adult injuries that kill:Repeated nerve injury  SA EAA toxicity  BDNF, GDNF, NT4 * **  trkB GAP43 p75  or  Schwann cell derived trophic factors • Immature state Regeneration mode

24. Adult injuries that kill: Avulsion Exogenous BDNF, GDNF prevents death No transport of Schwann cell derived NTFs EAA toxicity * **  trkB GFR1 p75 NOS No access to Schwann cells Regeneration possible  GAP43, CGRP, Tubulin Activation of p75 death pathway?

25. Rescuing dying motoneurons Preventing excitotoxicity • Riluzole • NOS inhibitors • Glutamate inhibitors MK801

27. Clinical trials in ALS • CNTFsevere side effects: fever, chest pains, muscle weakness, herpes virus activation • BDNF– Major side effects. Pain. • IGF1– data not conclusive; drug well tolerated • GDNF, NTN – not tested • TCH346 (anti apoptotic) – failed phase 2 trial • Riluzole– activity blocker • Retinoic acid– RALD2 & RAR

28. References • Lowrie MB & Vrbova G 1992 TINS 15: 80-84 • Greensmith L & Vrbova G 1995 Neuro-muscular Disord 5: 359-69 • Vejsada R et al 1995: EJN 7:108-115 • Ma J et al. 2001 139: 216-223