An investigation of the sensory and motor innervation of extraocular muscles in monkey and rat with combined tract-tracing and immunofluorescence methods: evidence for a dual motor innervation as common concept in mammals

The oculomotor system is one of the best studied motor systems.
Afferents from a variety of premotor areas converge on the
motoneurons in the three oculomotor nuclei to produce the different
types of eye movements. All oculomotor motoneurons participate in
all types of eye movements, and it was generally accepted, that
these motoneurons form a relative homogenous group which provides
the final common pathway for extraocular muscle (EOM)-motor
innervation. The EOM in mammals, the effector organs of the
oculomotor system, are fundamentally different from skeletal
muscle. They have two functionally different layers, global and
orbital layer, and are composed of two major muscle fibre classes,
singly-innervated (SIF) and multiply innervated fibres (MIF).
Previous studies in monkey revealed that SIF and MIF motoneurons
are anatomically separated and have different premotor inputs,
which support the idea of a dual motor innervation of EOM rather
than a final common pathway from motoneuron to EOM. Up to date,
neither motoneuron type has been further characterized nor has any
study proven their presence in other species to support the
hypothesis of the dual motor innervation as a common concept in
mammals. The functional implication of this system remains
speculative, though a role of MIFs together with their motoneurons
in a sensory feedback system controlling the EOMs is quite possible
and heavily debated. However, the lack of a common proprioceptor in
eye muscles does not support this theory. In monkeys SIF and MIF
motoneurons of extraocular muscles were identified by tracer
injections into the belly or the distal myotendinous junction of
the medial or lateral rectus muscle and further characterized by
combined tracer detection and immunohistochemical methods. The
experiments revealed that the MIF motoneurons in the periphery of
the motor nuclei lack non-phosphorylated neurofilaments,
parvalbumin and perineuronal nets, whereas SIF motoneurons
intensively express all three markers. In addition to the
histochemical differences, the MIF motoneurons are on average
significantly smaller in size than the SIF motoneurons. Analogous
to the study in monkey, the SIF and MIF motoneurons of the medial
and lateral rectus muscle of rats were identified with tracer
injections and further characterized by immunolabelling. For the
first time it was shown that both motoneurons types are present in
rat as well. The MIF motoneurons lie mainly separated from the SIF
motoneurons, and are different in size and histochemical
properties. As in monkey, the smaller MIF motoneurons lack
non-phosphorylated neurofilaments and perineuronal nets, both of
which are definite markers for the larger SIF motoneurons. A
possible proprioceptive control of eye movements requires the
presence of proprioceptive structures. The palisade endings
represent the best candidate for an EOM-proprioceptor. They were
analysed using antibody stains against the synaptosomal associated
protein of 25kDA, SNAP-25. With this robust method palisade
ending-like structures were identified for the first time in the
extraocular muscles of the rat. Furthermore the rat palisade
endings show characteristics of sensory structures thereby
supporting their role in proprioception. In conclusion, the EOM of
both monkey and rat are innervated by two sets of motoneurons which
differ in localization, morphology and molecular components. These
findings further support the presence of a dual motor control of
EOM that may apply widely to mammals, since it was verified in
monkey and rat. Palisade endings are a ubiquitous feature of mammal
EOM and most likely provide sensory information used for the
proprioceptive control of eye movements.