| dc.description.abstract | The effects of post-synaptic membrane blockade in the neuromuscular
junctions have been investigated in the developing chick embryo. Decamethonium
bromide (DeB), a non competitive blocker of acetylcholine that binds strongly to the
postsynaptic membrane of the neuromuscular junctions was applied at 0.2% solution
in normal saline on the chorioallantoic membrane for the experimental group while
the controls received normal saline.
Body weights, skeletal muscle histology and ultrastructure were compared
between age-matched control and experimental embryos from day 7 to day 20 of
incubation. Bone histology of the tibia and the general ossification pattern was
similarly observed from day 7 to 14.
Immunohistochemical fibre reactions were carried out on serial cryostat
sections from control and experimental embryo leg muscles associated with the tibia.
The sections were stained with F59 antibody, which stains myosin heavy chains in fast
fibres, S46 antibody which stains myosin heavy chain in slow fibres and anti desmin
which stains desmin at the periphery of the Z-line of muscle fibres.
The control and experimental embryo muscles were also studied by Electron
and Light microscopy to show both ultrastructure and general histology respectively.
Serial sections from the tibia were stained with Masson's trichrome, Alcian blue and
Hematoxylin to show bone formation, cartilage and general structure respectively.
Alizarin red and Alcian blue staining was performed on whole embryos to determine
cartilage and bone development.
The experimental embryos showed massive subcutaneous oedema, reduced
body weights and high mortality compared with the controls, which never had any
oedema. Muscle development was retarded and immunohistochemical reaction
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revealed delayed muscle patterning as well as degeneration in the experimental
embryos. There was almost complete skeletal muscle degeneration in the experimental
group of embryos by 14th day of incubation.
Electron microscopy showed well-developed myofibrils in the controls while
the same did not form in the experimental group. In both the control and experimental
embryos, the muscle membrane appeared intact however, the cytoplasm was dense
and vacuolated in the experimental group. Histological sections showed that most of
the degeneration process in the muscles in the experimental embryos started after the
application of second dose of DCB.
Bone histology showed that timing and onset of ossification of the tibia was
not affected by postsynaptic blockade in the experimental embryos although the size
of tibia was reduced by about 30% in comparison to the controls. There was fusion of
the intervertebral joints in the cervical region and also the femorotibial joint. The
lowerjaw was found to protrude beyond the upper jaw in the experimental embryos.
The body weights of the control embryos were found to be significantly higher
(p<0.05) than in the experimental group and this was more pronounced from day 16 of
incubation.
Decamethonium bromide induced blockade produced complete paralysis in the
experimental embryos and interfered with normal myosin heavy chain formation,
which leads to disturbed muscle patterning and degeneration. The degenerative
changes in muscles in turn resulted in subcutaneous fluid accumulation due to reduced
muscle contraction and slowed venous return. Paralysis in the embryos resulted in
poor joint formation and in some cases, joint fusion due to lack of skeletal movements
around the articular surfaces. On the other hand DCB did not to have any influence
on the ossification process of the long bones, which appears to be under an
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independent genetic program. Skeletal muscle paralysis caused reduced long bone size
by 30%. Bone size development is affected by the intermittent cycles of compression
and tension from muscles, which leads to calcification, remodelling and shaping of the
bones. Paralysis counteracted the beneficial effect of muscle contraction thereby
leading to lower bone sizes in the experimental embryos.
In conclusion, DCB induced synaptic blockade caused severe skeletal muscle
degeneration, disturbed muscle patterning, joint fusion between bones and reduced
size of the long bones in the chick embryo. However DCB did not affect the timing of
bone ossification. | en |
