A Subset of Palisade Endings Only in the Medial and Inferior Rectus Muscle in Monkey Contain Calretinin

Abstract

PurposeTo further chemically characterize palisade endings in extraocular muscles in rhesus monkeys.MethodsExtraocular muscles of three rhesus monkeys were studied for expression of the calcium-binding protein calretinin (CR) in palisade endings and multiple endings. The complete innervation was visualized with antibodies against the synaptosomal-associated protein of 25 kDa and combined with immunofluorescence for CR. Six rhesus monkeys received tracer injections of choleratoxin subunit B or wheat germ agglutinin into either the belly or distal myotendinous junction of the medial or inferior rectus muscle to allow retrograde tracing in the C-group of the oculomotor nucleus. Double-immunofluorescence methods were used to study the CR content in retrogradely labeled neurons in the C-group.ResultsA subgroup of palisade and multiple endings was found to express CR, only in the medial and inferior rectus muscle. In contrast, the en plaque endings lacked CR. Accordingly, within the tracer-labeled neurons of the C-group, a subgroup expressed CR.ConclusionsThe study indicates that two different neuron populations targeting nontwitch muscle fibers are present within the C-group for inferior rectus and medial rectus, respectively, one expressing CR, one lacking CR. It is possible that the CR-negative neurons represent the basic population for all extraocular muscles, whereas the CR-positive neurons giving rise to CR-positive palisade endings represent a specialized, perhaps more excitable type of nerve ending in the medial and inferior rectus muscles, being more active in vergence. The malfunction of this CR-positive population of neurons that target nontwitch muscle fibers could play a significant role in strabismus.

Palisade Endings Have an Exocytotic Machinery But Lack Acetylcholine Receptors and Distinct Acetylcholinesterase Activity

“…Consistent with previous studies, 21 , 33 , 34 nerve fibers forming palisade endings exhibited ChAT immunoreactivity. We found palisade endings which were supplied by single axons and others by up to four axons ( Figs.…”

“… 10 , 11 , 21 By immunolabeling, we demonstrated that all nerve fibers forming palisade endings expressed ChAT and all terminal varicosities of palisade endings expressed ChAT as well. These findings confirmed previous studies 17 , 21 , 34 that showed that palisade endings are exclusively cholinergic.…”

“… 39 At the molecular level, we identified key proteins of the exocytotic machinery in palisade endings including SNAP25, synaptobrevin, syntaxin, synaptotagmin, and complexin. As previously shown in monkeys, 34 SNAP25 was enriched throughout the palisade endings and was also present along axons supplying palisade endings albeit at lower concentration. The same distribution pattern existed for syntaxin, and complexin, whereas synaptobrevin and synaptotagmin were located in restricted areas of palisade endings.…”

“… 8 , 35 Further, in two primate species (man and monkey) many palisade endings located in the medial rectus muscle and a lower number of palisade endings of the inferior rectus muscle express calretinin, whereas palisade endings of the other two rectus muscles lack calretinin. 19 , 34 Calretinin is a calcium-blinding protein and, although its functional role is unclear, the authors have hypothesized that calretinin-positive palisade endings represent a specialized, probable more excitable, type of palisade ending needed for convergent eye movement. 19 , 34 Recently, it has been demonstrated that MIF motoneurons participate in the whole repertoire of eye movements.…”

“… 19 , 34 Calretinin is a calcium-blinding protein and, although its functional role is unclear, the authors have hypothesized that calretinin-positive palisade endings represent a specialized, probable more excitable, type of palisade ending needed for convergent eye movement. 19 , 34 Recently, it has been demonstrated that MIF motoneurons participate in the whole repertoire of eye movements. 49 Due to the fact that palisade endings are extensions of MIF motoneurons axons, this suggests that palisade endings despite their unknown function, are relevant for all types of eye movement.…”