Pumps, Channels and Membranes

Transcription

1 Pumps, Channels and Membranes A bit of heresy. PLEASE read Alberts et al. or equivalent, for counterpoint

2 Current view Cell K + K channel Na + Na channel

3 How did these concepts arise? membrane (~200 years ago) semi-permeable (passes water only) K + leakage: K + channel (atomic sieve) Na + leakage! Na + channel (+K + channel) Na + accumulation in cell? Na + -pump req d to prevent K + loss from cell? K + -pump req d additional pumps and channels

4 Reflections Pumps and channels arose out of ad hoc hypotheses Most/all solutes partition out of electrochemical equilibrium therefore, all of these solutes require pumps, channels Number of putative pumps/channels > 100 Many membrane-containing widgets: huge complexity.

5 Do channels and pumps really exist?

6 Channel considerations Patch-clamp experiments Dog-door conundrum

7 Patch clamp experiment:

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11 Issue #2: Large channels and small solutes. membrane membrane

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13 Pump issues Enough energy to drive the pumps? Enough space to accommodate them?

14 Membrane pumps: require energy Na-pump consumes 30-35% of cell s energy supply 50+ additional membrane pumps pumps in mitochondrial membrane pumps in endoplasmic reticular membrane etc. Is there enough energy?

15 Poison-cocktail experiment Cyanide, iodoacetate, nitrogen: cut off energy Ion gradients maintained for eight hours Compute amount of pumping required Compare available energy RESULT: shortfall times (conservative) Therefore, not enough energy even for Na pump

16 next issue: Space for pumps (and channels)? New drugs continue to emerge New drug implies new pump Will a limit be reached?.

17 Summary of concerns Channels: patch clamp selectivity (dog door problem) Pumps: energy space

18 To here 09

19 Could the root problem be more foundational?

20 Train of logic: Membrane assumed continuous and impermeant gave rise to channels, pumps channels, pumps questionable Alternative: membrane discontinuous, or permeant Channels and pumps perhaps unnecessary

21 If membrane discontinuous/permeant, then disruption should be relatively innocuous. After all, what s another hole? What happens when membrane is disrupted?

22 What happens when large hole is punched?

23 Should be no problem

24 Cut cell experiment: Should be no problem

25 Conclusion: Membrane continuity apparently unnecessary for life, function (many examples)

26 Possibility: membrane not really continuous, after all (Creating additional holes will make little difference) Evidence for absence of membrane continuity?

27 Membrane contains much protein: natural portals

28 Membrane wounding: Creates holes through which large molecules that cannot ordinarily pass do pass

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30 Thus, lipid bilayers do appear to be leaky, or discontinuous (Lots of proteins, and possibly physical discontinuities)

31 Could explain how cells change volume. Lipid bilayer can t accommodate. Proteins can unfold Small volume Large volume

32 side questions re: membrane anomalies Note also: removal of phospholipid does not alter EM image.

33 Therefore: lingering questions about membrane appears uninterrupted, but where are the proteins? persists in EM after phospholipid is removed bacteria: appears in EM, despite no fatty acids What s going on? -- unclear

34 Conclusion Membrane appears to be present (?) Continuity of impermeable barrier unlikely.

35 Is the root problem still deeper?

36 Why was continuous barrier postulated to begin with? to maintain cellular integrity; to prevent mixing btw. inside and outside of cell -cytoplasm assumed to be aqueous solution Is the cytoplasm really an aqueous solution?

37 Sure seems like gel!

38 Cytoplasm: Aqueous solution or gel? contains charged polymers (30%); water (70%) polymers cross-linked, as in typical gel To confirm gel-like consistency: Look at behavior of water, ions

39 Cell water freezing temperature appreciably depressed trees plants cold-blooded animals Interpretation: water molecules constrained; can t enter ice configuration.

40 Ions: freely diffusible?

41 1. Static localization:

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43 from Edelmann, 1989

44 2. Dynamic localization:

45 Conclusions Water molecules constrained (around surfaces) Ions constrained onto surfaces Therefore: NOT an aqueous solution Similar to artificial gels

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48 End of session 2