The source of innervation and function of palisade endings in the extraocular muscles of Macaca mulatta

Vertebrate extraocular muscles show a highly complex anatomy, which
differs in many respects from skeletal muscles. Furthermore, there
is a considerable variation among different species with regard to
the presence of proprioceptive organs. Whereas muscle spindles and
Golgi tendon organs are well developed in sheep and pig, neither
are found in cat, and only poorly developed muscle spindles are
present in human and monkey. In all vertebrates studied so far
cuffs of nerve terminals around multiply innervated muscle fibers
of the global layer, termed palisade endings (PE) are present at
the myotendinous junction. Palisade endings (PE) are specialized
nerve endings unique to extraocular muscles. There is still an
ongoing debate on PE function. A proprioceptive function is
supported by their ultrastructural morphology, and the location of
the majority of their terminals within the muscle tendon. A motor
function of PEs is suggested by the expression of different
cholinergic markers, and the binding of α-bungarotoxin to at least
a small proportion of PE nerve endings. So far the location of the
somata giving rise to the PEs is unknown. After eye muscle
injections with different tracers, and the investigation of
retrogradely labeled cells after three days of survival, there are
two possible options for the location of the cell bodies of PEs:
The trigeminal ganglion (TG), where almost all of the sensory
afferents of eye muscles come from, or the motor neurons of the
abducens (nVI), trochlear (nIV) and oculomotor nucleus (nIII) in
the brainstem, where the motor neurons of the extraocular muscles
are situated. In the first part of this thesis the histochemical
and morphometric properties of TG cells projecting to the
extraocular muscles were studied and related to different nerve
fibers and terminals in these muscles, to obtain more information
about their function. Retrogradely labeled TG neurons and eye
muscle terminals were processed for the presence of substance P
(SP), nitric oxide synthase (NOS), calretinin (CR) and
cholinacetyltransferase (ChAT). Injections of the tracer were
placed into the medial rectus (MR) or the lateral recuts muscle
(LR), either in the belly or in the distal, myotendinous part of
the muscle. PEs were only labeled by CR or ChAT and none of the
other markers that were used. Furthermore, CR positive retrogradely
labeled tracer cells in the TG were rarely found (under 1 % of all
tracer cells). The results indicate that the possible function of
TG afferents could be mainly vasodilatation or nociception whereas
the probability of proprioception via PEs is very low. These
results point to the fact, that the source of the PE cell body is
probably localized in the brainstem. In a second attempt to solve
this problem, rhesus monkey received tract-tracer injections (WGA
or CT) into the oculomotor nucleus, which contains the motor
neurons of the medial, inferior and superior recti and inferior
oblique muscles, as well as the trochlear nucleus, which contains
the motor neurons of the superior oblique. All extraocular muscles
were processed for the combined immunocytochemical detection of the
tracer and non-phosphorylated neurofilament for the visualization
of the complete muscle innervation. In all muscles (except lateral
rectus) numerous anterogradely tracer labeled PEs were found, as
well as tracer-filled tendon organs. In addition the en plaque and
en grappe motor endings, were also strongly tracer positive. Double
immunolabeling revealed that all types of nerve endings including
tendon organs were anterogradely labeled, except the thin tyrosine
hydroxylase positive autonomic nerve fibers of the sympathetic
system. No anterograde labeling was found within the trigeminal
ganglia. These results suggest that the somata of palisade endings
are located within the brainstem, in or around the oculomotor
nuclei, and confirm several previous studies. In how far the
multiple nerve endings of non-twitch muscle fibers and the PEs form
an anatomical entity with one parent soma in the periphery of the
oculomotor nuclei was studied by morphological and histochemical
analysis of the peripheral neurons around the motor nuclei. This
data revealed two populations: one group of cholinergic multipolar
neurons represent the motor neurons supplying the multiple motor
innervation, one group of round calretinin positive cholinergic
neurons giving rise to PEs. If the palisade endings do have a
sensory function, then their cell body location amongst the
non-twitch motor neurons would be an ideal location to control the
tension in the non-twitch extraocular muscle fibers, and the data
brought together in this thesis points strongly in this direction.
To fully establish this hypothesis more physiological experimental
data is required.