Glutamatergic modulation of synaptic‐like vesicle recycling in mechanosensory lanceolate nerve terminals of mammalian hair follicles
Authors: Banks Robert W. , Cahusac Peter M. B. , Graca Anna , Kain Nakul , Shenton Fiona , Singh Paramjeet , Njå Arild , Simon Anna , Watson Sonia , Slater Clarke R. , Bewick Guy S. 
Published: February 2013

Key points
The lanceolate sensory nerve ending of hair follicles is known to contain small (∼50 nm), clear vesicles similar to those of presynaptic terminals, but of unknown function.
We show that the sensory terminals spontaneously take up and release the fluorescent styryl dye FM1‐43, and also provide other evidence that the dye flux is primarily by recycling of these synaptic‐like vesicles (SLVs).
FM1‐43 labelling is Ca2+ dependent, and its release is sensitive to α‐latrotoxin, which is known to deplete synaptic vesicles at neuromuscular junctions.
Responses of hair follicle afferents are not significantly affected by FM1‐43 at a concentration (10 μm) sufficient to label the endings, so the mechanotransduction channel that has previously been shown to be blocked by FM1‐43 permeation in hair cells of the inner ear and in cultured dorsal root ganglion cells is either not responsible for sensory transduction in the lanceolate ending or is in some way protected from exposure to the dye.
The sensory terminals are relatively enriched in glutamate, presumably within the vesicles.
Exogenous glutamate increases FM1‐43 labelling, whereas the labelling is strongly inhibited by PCCG‐13, a specific blocker of a non‐canonical phospholipase D‐linked metabotropic glutamate receptor, but not by canonical ionotropic or metabotropic glutamate receptor blockers. It is also inhibited by FIPI, a novel phospholipase D inhibitor.
The system of SLVs is closely similar to that we have previously described in the muscle spindle, and where we further demonstrated the regulatory action of glutamate on the sensory response to maintained stretch.
We conclude that an SLV‐mediated glutamatergic system is present in the mechanosensory endings of the primary afferents of lanceolate endings, and it appears to function in a similar way to the autoregulatory system of the muscle spindle.
Abstract  Our aim in the present study was to determine whether a glutamatergic modulatory system involving synaptic‐like vesicles (SLVs) is present in the lanceolate ending of the mouse and rat hair follicle and, if so, to assess its similarity to that of the rat muscle spindle annulospiral ending we have described previously. Both types of endings are formed by the peripheral sensory terminals of primary mechanosensory dorsal root ganglion cells, so the presence of such a system in the lanceolate ending would provide support for our hypothesis that it is a general property of fundamental importance to the regulation of the responsiveness of the broad class of primary mechanosensory endings. We show not only that an SLV‐based system is present in lanceolate endings, but also that there are clear parallels between its operation in the two types of mechanosensory endings. In particular, we demonstrate that, as in the muscle spindle: (i) FM1‐43 labels the sensory terminals of the lanceolate ending, rather than the closely associated accessory (glial) cells; (ii) the dye enters and leaves the terminals primarily by SLV recycling; (iii) the dye does not block the electrical response to mechanical stimulation, in contrast to its effect on the hair cell and dorsal root ganglion cells in culture; (iv) SLV recycling is Ca2+ sensitive; and (v) the sensory terminals are enriched in glutamate. Thus, in the lanceolate sensory ending SLV recycling is itself regulated, at least in part, by glutamate acting through a phospholipase D‐coupled metabotropic glutamate receptor.