Unexpected Role for Laminin in Skin Sensation
In the skin, sensory neurons are in charge of detecting external stimuli, but they do not go it alone. A new line of research shows that skin cells, and the extracellular matrix proteins they produce, may be important modulators of pain sensation. A study published online July 3 in Nature Neuroscience, from Gary Lewin, Max Delbrück Center for Molecular Medicine, Berlin, Germany, and Jing Hu, Center for Integrative Neuroscience, Tübingen, Germany, reveals that keratinocytes in the epidermis produce a unique extracellular matrix that inhibits the responsiveness and branching of mechanosensitive neurites, including those responsible for pain. The researchers pinpointed the matrix component laminin-332 as the factor responsible for the inhibition.
Besides revealing a new role for the matrix in corralling sensory neuron growth and activity, the study offers potential insight into the painful skin disease epidermolysis bullosa, which is caused by loss of laminin-332 expression.
In the new study, first authors Li-Yang Chiang and Kate Poole, and their coworkers, used whole-cell patch clamp techniques to measure mechanosensitive currents in dorsal root ganglion (DRG) sensory neurons in culture. They found that the laminin matrix produced by mouse keratinocytes, but not that from other kinds of cells, suppressed one type of mechanosensitive current, the rapidly adapting (RA) current. Further studies tracked that inhibition to laminin-332 (formerly called laminin-5), an isoform specific to keratinocyte-derived matrix. In skin, keratinocytes reside in the outer layer, the epidermis, which is innervated by nociceptors. Therefore, these results raise the possibility that laminin-332 dampens the response of epidermal nociceptors, acting as a brake on painful sensations.
Mechanotransduction is mediated via poorly understood ion channels, and there are two views about how the channels are opened. One view holds that stretching the neuronal membrane results in channel opening. However, Lewin told PRF that there is also evidence that mechanosensory channels are linked to the extracellular matrix and open when force is transferred to them from the matrix. Previously, Lewin, Hu, and their colleagues reported that the appearance of mechanoresponsive currents in DRG sensory neurons (and some nociceptors) requires a protein filament that tethers neurons to the matrix (Hu et al., 2010).
The group wondered whether that tether might be missing in the keratinocyte matrix. Indeed it was: Transmission electron microscopy (TEM) of sensory neurons cultured on laminin showed that when the matrix contained laminin-332, the tethers disappeared.
To figure out how laminin-332 might be having its inhibitory effect, the researchers plated sensory neurons on a surface micro-printed with a laminin grid. Strikingly, RA mechanosensitive currents could be evoked from neurites growing on laminin-332–free stripes, but neurites sitting on neighboring laminin-332 stripes rarely showed those currents. That was true even for neurites in the same cell. Because stripes were only 25 μm apart—a tiny space compared to the area covered by a neuron’s receptive field—the authors conclude that laminin-332 acts on neurites locally, presumably disrupting attachment of the tether to the matrix and thus preventing ion channels from opening.
In addition, neurites grew preferentially on stripes lacking laminin-332. Growth cones regularly branched on the permissive matrix, but rarely sent new neurites into matrix containing laminin-332.
Lewin wonders whether neurites pick their paths by actually “feeling” the shape of the laminin matrix, which atomic force microscopy (AFM) showed became bumpier upon addition of laminin-332. “It could be that the growth cone is sensing mechanical force, like your finger does if you run it over sandpaper,” he said.
In addition to their studies on cultured mouse cells, the team investigated tissue from patients with Herlitz-type junctional epidermolysis bullosa (EB), a skin disease caused by mutations in the genes for any of laminin-332’s three subunits. The lack of laminin-332 leads to extreme skin blistering and pain, and this type of EB is often lethal in the first year of life. The researchers found that laminin matrix from keratinocytes of healthy people dampened RA mechanosensitive currents in cultured neurons, whereas matrix from patients with EB failed to suppress those currents. Further, skin biopsies showed changes in nerve branching: In healthy skin, laminin-332 was present at the junction between dermis and epidermis, and sensory fibers that grew across that border were usually unbranched. In patients with EB, fibers branched at the border.
The observed increases in neuron excitability and branching may help to explain the pain that marks EB. The findings may also shed light on how sensory neurons are regulated normally. Previously, Lewin says, laminin-332 was seen strictly as a structural protein that helps to maintain skin integrity; now, his group’s findings reveal that it also holds sway over sensory neurons. For neurons that lie just below the skin’s surface, he speculates, laminin-332 might create “an endogenous inhibition mechanism…to make sure those fibers are not oversensitive.”
This study joins a number of other new findings on the role of keratinocytes in modulating sensory neurons and pain. Recently, researchers from the labs of Phillip Albrecht and Frank Rice at Albany Medical College and Integrated Tissue Dynamics, Rensselaer, New York, reported that calcitonin gene-related peptide β (CGRPβ) from keratinocytes—previously thought to be CGRPα from neurons—may drive a variety of chronic pain conditions (Hou et al., 2011).
Finally, Lewin’s study hints that the extracellular matrix may constitute a new target for treating peripheral pain. In the case of laminin-332, Lewin says that small molecules mimicking its action, applied topically, could possibly serve as analgesic agents for a wide range of pain conditions. He notes that, in addition to EB, other diseases such as diabetic neuropathy involve changes in epidermal innervation.
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