Determination of Threshold Levels of Sour Gas and H 2 S on the Mammalian Brain

1. Determination of Threshold Levels of Sour Gas and H2S on the Mammalian Brain Funding Agreement Number 2001-01 BC Oil and Gas Commission Sheldon H. Roth Faculty of Medicine University of Calgary

2. Objectives of Study • to determine threshold levels of hydrogen sulphide (H2S), sour gas and H2S/organic compound mixtures i.e. the Lowest Observable Adverse Effect Levels (LOAELs) and the No Observable Adverse Effect Levels (NOAELs) • to evaluate the differences of effects between H2S alone and as a component of sour gas on in vivo and in vitro mammalian central nervous systems (CNS)

3. Hydrogen Sulphide (H2S) • extremely toxic gas • environmental & industrial pollutant • produced by variety of human and natural processes • produced endogenously in mammals

4. Hydrogen Sulphide • colourless • heavier than air • flammable • characteristic “rotten egg” odour • odor threshold 0.5 ppb • olfactory fatigue > 100 ppm ATSDR 1999 H2S Health Research & Risk Assessment Symposium 2000

5. H2S exposure • “one of the leading causes of sudden accidental death in the workplace” • affects many organ systems (broad spectrum toxicant) • effects dependent on concentration, duration and rate of exposure • organs with high O2 demand - most susceptible

6. Broad spectrum effects of H2S • nervous system (brain) • lung • heart • reproductive & developing systems • eye • metabolic systems • smell (olfactory)

7. Dose Response of H2S Roth Env Epidemiol Toxicol 1999

8. Effects of H2S (low level exposure) • eye, nose & throat irritation • headache, sinus, etc • nausea • hoarseness, cough • nasal congestion • shortness of breath • stress • drowsiness Schiffman H2S Health Research & Risk Assessment Symposium 2000

9. Effects of H2S exposure on the nervous system (brain) • mental depression • irritability • loss of memory • lethargy • dizziness • loss of appetite • fatigue • headache memory Fundamentals of Neuroscience 1999

10. hippocampus • functions of the hippocampus • memory storage • cognitive function • sensory processing • voluntary movements Amaral & Witter, 1989

11. Recording from hippocampus placement of stimulating & recording electrodes PS = amplitude of population spike

12. Methods • prepare rat hippocampal slices • place slices in recording chamber • activate excitatory synaptic inputs to CA1 pyramidal neurons • record field potentials (PS) of CA1 neurons • apply H2S gas (in nitrogen) • analyze data using custom software

13. Monitor Interscan Corporation 4000 Series Compact Portable Analyzer 0-1999 ppb 1 ppb resolution electrochemical voltammetry sensor

14. control experiments nitrogen

15. Exposure to 50 ppm H2S

16. Exposure to 25 ppm H2S 25 ppm H2S

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18. Exposure to 1 ppm H2S 1 ppm H2S

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20. 500 ppb H2S

21. Dose response curve - H2S

22. Inhibition of Respiratory Mechanisms cytochrome c oxidase whole brain IC50 = 0.13 µM Nicholson et al J Toxicol Environ Health A 1998

23. J Toxicol Environ Health A 1998

24. Conclusions • long-lasting synaptic enhancement occurs following exposure to low levels of H2S • effects appear to be concentration-dependent • inhibition of respiratory enzymes (cytochrome oxidase, carbonic anhydase) confirmed • effects may account for symptoms such as anxiety and cognitive impairment

25. Investigators Ron Bennington James Brooks Russell Nicholson (SFU) Lisa Partlo R.S. Sainsbury Barbara Skrajny Lisa Strosher Steven Ip Ricardo Bonafine Peter O’Brien (U of T) D.P. Archer Funding BC Oil & Gas Commission Medical Research Council of Canada Alberta Heritage for Medical Research University of Calgary Acknowledgements

26. Future Experiments • H2S experiments in ppb range • S. Ip • Sour gas experiments • S. Ip • whole animal (in vivo) studies • Dr. R. Bonafine • further studies on respiratory enzymes • Dr. P. O’Brien University of Toronto

27. Sour Gas

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