Vacuole transport lab, and red beet tissue

Ross Koning Koning at ECSUC.CTSTATEU.EDU
Mon Nov 11 18:11:54 EST 1996


At 10:45 AM 11/9/96 -0800, Scott T. Meissner wrote:
>While studying the water potential of red beet storage
>tissue I found that this tissue is isotonic with from
>0.2 to 0.4 M sorbitol, not water.  Amazed the heck out
>of me since I got the idea previously that since the
>plant takes up ground water this fixes the base water
>potential of its organs.

I find most root "sap" (ground up root tissue, centrifuged)
runs about 0.3-0.4 osmolar.  So surely roots are turgid
since soil water is very close to 0.0 osmolar (in most
farmland soils).  I never expect roots to be at equilibrium
with soil water...plants survive on gradients, not equilibria.
It is important to separate osmotic potential from water
potential and to not assume equilibrium conditions.

>Now, if you take a one cell layer thick section of beet
>tissue, all the cells will be exposed to the solution they
>are placed in.  Under these conditions you can see the
>swelling and lysis of many of the cells over time.  The
>vacuole stretches and bursts into the cytosol, I have seen
>this happen on many of the times I have been trying to
>measure the incipient plasmolysis point of this storage
>tissue.  So with a one cell thick layer the cells will
>all suffer high turgor.

I think all the root cells develop high turgor...surely by the
end of a 30 min incubation most of the cells are **nearing**
equilibrium.  In a thick plug, admittedly cells in the
interior are responding more slowly as there is a trapped
apoplast.  Our plugs are about 5 cm diam.  I think that
having some thickness is the saving grace rather than the
condemnation of using plugs (see below).

On the other hand, a one-cell-thick slice has some other
difficulties, in my opinion.  First, a very high percentage
of the cells will have been damaged by the blade.  These
will leak because of that.  The method you described to
produce vacuoles demonstrates that surface effect.  Second,
I think these cells all have plasmodesmata...I wish I had
some stereological analysis of EMs of beet root cells.
In a one-cell thick slice these now represent membrane
areas not supported by the wall in holding back the
internal pressure.  It would not surprise me to see
cytoplasmic "blebs" and considerable leakage of these
cells in distilled water (soil water).

In this way I see the plug as better-representing what a root
does than a thin slice...too much artifact (surface damage
effects) in the latter.

>My main point was that the beet storage tissue is not isotonic with
>water.

I agree with you on this.

>Taking this tissue and sticking it in water will result in
>extremely high turgor pressures in the exposed cells, damaging most
>of them to the point of lysis.

I just have never observed that after a rinse of perhaps a few
minutes.

>So following the movement of betacyanins
>out of a beet tissue plug that is placed in water may not reflect
>membrane transport processes as much as cell death.

See, in my hands the betacyanins just don't **do** that.
I get betacyanin release in solutions OTHER THAN water.

>This experiment
>needs to be run under conditions that are isotonic to the storage
>tissue, and when it is then the cells will not be lysing due to
>high turgor and the leakage of betacyanins may be due to their
>crossing the membranes of living cells.

The control is distilled water...I get ***no*** betacyanin release.
The treatment is 4% NaCl...I get tremendous betacyanin release.
The control is 0.2% CaCl...I get no betacyanin release.
The treatment is 4% NaCl + 0.2% CaCl...I get very little betacyanin release.

With results like those, it is hard for me to argue that I
am seeing turgor-driven lysis of cells!  The Calcium ion
effect is impossible to explain invoking a pressure model.
I can only explain these results thinking about membrane
transport proteins.

ross

______________________________________________________________
                            |
Ross Koning                 | Koning at ecsu.ctstateu.edu
Biology Department          | http://koning.ecsu.ctstateu.edu/
Eastern CT State University | Phone: 860-465-5327
Willimantic, CT  06226  USA | Fax: 860-465-5213
____________________________|_________________________________

                Plant Physiology is Phun!

 /\|___/\     //\______COOH   NH-CH2-CH=C-CH2OH  \/OH
|  |  |  |    |  |  ||       //\___     \CH3     /\|/\\/\\COOH
 \/ \/|\/|    \\/ \ /       N  ||  N            |  |
 /\ | |__|=        NH       |  || ||           //\//\
  | COOH                    \\ /\ /            O
  COOH        H2C=CH2         N  NH
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