a selection of unanswered questions

Terence P. Ma tpm at netdoor.com
Wed Jun 5 09:03:57 EST 1996

Margaret Fowler wrote:

>        The following questions have never been answered satisfactorily,
> several of them never at all:-

Many of these questions are not in my field of expertise. However, a number 
of these issues are addressed extrememly well in several books and 
textbooks, in addition to journal articles. I am surprised that people have 
answered these inquiries, asked in a most aggressive and combative format, 
at all. If serious discussion were desired, I assume that appropriate 
citations for some of these questions would have been provided. Since I 
don't have all my citations at hand, I won't give all of them, but will be 
rather general.

> Question 12: How can one study membranes by electron microscopy, when
>     they are believed to contain lipids which the procedure extracts?

Addressed extensively in Hyatt's textbook on electron microscopy and 
practically any decent textbook on EM.
> Question 13:  What is the real evidence that rapid deep freezing for
>     electron microscopy causes less shrinkage and distortion of tis-
>     sues, cells and organelles, than classical transmission electron
>     microscopy?

See another of Hyatt's texts and see Peters, Palay, and Webster.

> Question 14:  Why do those who calculate dimensions from electron
>     micrographs not take into account the shrinkage during preparation
>     and examination of their sections, cells and organelles?

See above. And who says we don't in some fashion. Read the Materials and 
Methods sections more carefully. I often discuss shrinkage artifact in my 
> Question 17:  Why should it be necessary to tilt the stage of the
>     electron microscope to see randomly orientated membranes in all
>     orientations, when this is not necessary with the light microscope?

See Hyatt. Look in any basic physics textbook re: incidence of light and 
electron beams.

> Question 21: Why are receptors and channels, which have been character-
>     ised, sequenced and their sizes measured or calculated, not seen
>     on membranes by transmission electron microscopy?

See Peters, Palay, and Webster. I believe they show at least one example.

> Question 28: Why are the dimensions and numbers of synapses
>     different by light and electron microscopy?

Who says they are?
> Question 29:  Why are there no light micrographs in the literature
>     showing the connection of one cell body by a dendritic pre-
>     synaptic fibre to a synapse on another cell body?

There aren't. Guess the ones in my papers don't count. Of course, one never 
knows for a fact that a presumptive contact is a real contact unless one 
does the correlative EM. And THAT has been done in a large number of papers 
dating back to the 1970s.
> Question 36:  In immunocytochemistry, is it assumed that the fixatives,
>     dehydrating reagents, washings, and primary and secondary anti-
>     bodies, do not change the reaction of the antibody to the antigen
>     believed to be in a particular cell or part of a cell?

No. See Cuello's book on Immunohistochemistry.

> Question 37:  Is it reasonable to believe that processes or dendrites
>     contain different antigens from the cell bodies from which they
>     arise?

Question currently being debated and investigated. Read the past five years 
of Journal of Comparative Neurology for an idea of the papers covering this 
> Question 39:  Is it warrantable to assume that growth of tissues in
>     culture does not change their morphology, biochemistry, or
>     immuno-reactivity?

Sure changes morphology. Many people devotes years to that study. Its called 
study of development (on one end) and aging (on the other). Much print has 
been devoted to this. See Purves and Lichtman's or Max Cowan's text on 
development side. Lots of review articles on the aging side.

> Question 40:  Does not the use of the term neuroglia imply that the
>     authors can not distinguish between astrocytes, oligodendrocytes,
>     and microglia?

No. When I am write about neurons, I recognize that there are various types 
of neurons. Thus, why should I not use an all inclusive term for the various 
types of glia?

> Question 41:  Why are the individual types of neuroglial cells so
>     rarely seen by light microscopy of healthy central nervous systems?

Says who?
> Question 42:  Since the latter three alleged cell types were described
>     by classical histological techniques during the first half of the
>     twentieth century, does this not imply that anyone using anti-
>     bodies to mark them specifically must first identify them by
>     these criteria?

Why so?

> Question 48:  Why do synapses seen by electron microscopy appear so
>     much smaller than those seen by light microscopy?
Who says they are? They aren't. See reconstruction of synapse studies. See 
Peters, Palay, and Webseter.
                      Terence P. Ma, Ph.D.
Depts. of Anatomy and Neurology    Director of Officials
Univ. of Mississippi Med. Ctr.     Collegiate Water Polo Association
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Email:  tpm at Anat.UMsMed.Edu        Email:  tma at EWPRA.Org
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