Abstract
Merkel cell-neurite complexes are highly sensitive touch receptors comprising epidermal Merkel cells and sensory afferents. Based on morphological and molecular studies, Merkel cells are proposed to be mechanosensory cells that signal afferents via neurotransmission; however, functional studies testing this hypothesis in intact skin have produced conflicting results. To test this model in a simplified system, we asked whether purified Merkel cells are directly activated by mechanical stimulation. Cell shape was manipulated with anisotonic solution changes and responses were monitored by Ca2+ imaging with fura-2. We found that hypotonic-induced cell swelling, but not hypertonic solutions, triggered cytoplasmic Ca2+ transients. Several lines of evidence indicate that these signals arise from swelling-activated Ca2+-permeable ion channels. First, transients were reversibly abolished by chelating extracellular Ca2+, demonstrating a requirement for Ca2+ influx across the plasma membrane. Second, Ca2+ transients were initially observed near the plasma membrane in cytoplasmic processes. Third, voltage-activated Ca2+ channel (VACC) antagonists reduced transients by half, suggesting that swelling-activated channels depolarize plasma membranes to activate VACCs. Finally, emptying internal Ca2+ stores attenuated transients by 80%, suggesting Ca2+ release from stores augments swelling-activated Ca2+ signals. To identify candidate mechanotransduction channels, we used RT-PCR to amplify ion-channel transcripts whose pharmacological profiles matched those of hypotonic-evoked Ca2+ signals in Merkel cells. We found 11 amplicons, including PKD1, PKD2, and TRPC1, channels previously implicated in mechanotransduction in other cells. Collectively, these results directly demonstrate that Merkel cells are activated by hypotonic-evoked swelling, identify cellular signaling mechanisms that mediate these responses, and support the hypothesis that Merkel cells contribute to touch reception in the Merkel cell-neurite complex.
Highlights
The mechanical senses of touch, proprioception, hearing and balance are initiated by cells that transduce force into electrical signals
Our findings demonstrate that hypotonic stimuli cause Ca2+ influx in purified Merkel cells, and indicate that this Ca2+ influx is initiated by swelling-activated ion channels
If Merkel cells express ion channels activated by membrane tension, we reasoned that cell swelling might increase membrane tension to open such channels
Summary
The mechanical senses of touch, proprioception, hearing and balance are initiated by cells that transduce force into electrical signals. Though the mechanotransduction events underlying audition and balance have been extensively investigated [1,2], little is known about the signals that initiate the somatic mechanical senses of proprioception and touch [3,4]. Mechanotransduction occurs in peripheral afferent terminals, which are heterogeneous and widely distributed throughout the body, making most somatosensory mechanoreceptors difficult to study directly. Merkel cell-neurite complexes, which are cutaneous mechanoreceptors critical for fine shape and texture discrimination [4], have several characteristics that overcome these experimental challenges. Unlike most cutaneous touch receptors, the response patterns of these touch receptors have been identified in semiintact recording preparations: when the skin is displaced they generate the slowly adapting type I (SAI) response [5,6]. In transgenic Math1/nGFP mice, Merkel cells are labeled with green fluorescent protein (GFP) [7], permitting enrichment of Merkel cells from dissociated skin using fluorescence-activated cell sorting (FACS) [8]
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