Pumps, Channels and Membranes

A bit of heresy…….

PLEASE read Alberts et al. or equivalent,

for counterpoint

+

+

NaK

K channel

Na channel

Current view

Cell

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

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 .

Do channels and pumps really exist?

Channel considerations

• Patch-clamp experiments

• “Dog-door”

conundrum

Patch clamp experiment:

Large channels and small solutes

membrane membrane

Issue #2:

.

Pump issues

• Enough energy to drive the pumps?

• Enough space to accommodate them?

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?

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 15 -

30 times (conservative)•

Therefore, not enough energy even for Na pump

Space for pumps (and channels)?

• New drugs continue to emerge

• New drug implies new pump

• Will a limit be reached? .

next issue:

Summary of concerns

Channels:• patch clamp

• selectivity (dog door problem)

Pumps:• energy• space

To here 09

Could the root problem be more foundational?

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

If membrane discontinuous/permeant, then disruption should be relatively innocuous.

After all, what’s another hole?

What happens when membrane is disrupted?

What happens when large hole is punched?

Should be no problem

Cut cell experiment:

Should be no problem

Conclusion:

Membrane continuity apparently unnecessary

for life, function

(many examples)

Possibility: membrane not really continuous, after all

(Creating additional

holes will make little difference)

Evidence for absence of membrane continuity?

Membrane contains much protein: natural portals

Membrane wounding:

Creates holes through which large molecules that cannot ordinarily pass

do pass

Thus, lipid bilayers do appear

to be leaky, or discontinuous

(Lots of proteins, and possibly physical discontinuities)

Could explain how cells change volume. Lipid bilayer can’t accommodate. Proteins can unfold

Small volume

Large volume

side questions re: membrane anomalies

Note also: removal of phospholipid does not

alter EM image

.

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

Conclusion

• Membrane appears to be present (?)

• Continuity of impermeable barrier unlikely .

Is the root problem still deeper?

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?

Sure seems like gel!

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

Cell water

• freezing temperature appreciably depressed

•

trees•

plants•

cold-blooded animals

• Interpretation: water molecules constrained; can’t enter ice configuration.

Ions: freely diffusible?

1. Static localization:

from Edelmann, 1989

2. Dynamic localization:

Conclusions

• Water molecules constrained (around surfaces)

• Ions constrained onto surfaces

• Therefore: NOT an aqueous solution

• Similar to artificial gels

End of session 2