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Emerging Concepts in Ion Channel Biophysics
Wednesday Speaker Abstracts
15
Using Leeches to Discover Novel Ion Channels Involved in Mechanotransduction
Elizabeth Heath-Heckman
1
, Maurizio Pellegrino
1
, Diana Bautista
1
, Francisco F. De-Miguel
2
,
David Weisblat
1
.
1
University of California - Berkeley, Berkeley, CA, USA,
2
Universidad Nacional Autónoma de
México, Mexico City, Mexico, Mexico.
Mechanotransduction, mechanisms by which cells convert mechanical stimuli into electrical
activity, is a process conserved across all domains of life. Despite its importance,
mechanotransduction is not well understood at a molecular level. To better characterize the genes
involved, we used leeches in the genus
Hirudo
whose ventral nerve cord ganglia contain three
classes of mechanosensory neurons distinguished by their responses to light touch (T cells),
pressure (P), and potentially damaging stimuli (N). To determine which genes confer these
behaviors, we performed RNASeq of the above cell types and two non-mechanosensory portions
of the ganglion. Some of the most highly regulated transcripts correspond to ion channels already
implicated in mechanosensation, such as ASIC, Trp, and CNG. However, two hyperpolarization-
activated cyclic nucleotide-gated channels (HCNs) were also upregulated in P and N cells.
HCNs, while important in the etiology of chronic pain, have not yet been shown to be involved
in mechanosensation. The genome of
Helobdella robusta
encodes 7 HCNs in what appears to be
a lineage-specific gene amplification, suggesting that the roles of HCNs in leeches may differ
from, or be more specialized than, those in other animals. In situ hybridization showed at least
five of these HCNs are expressed in the nerve cord ganglia in juvenile animals, one of them
identical to the enrichment found in
Hirudo
. Preliminary experiments suggest that RNAi of the
upregulated HCN in
Hirudo
P cells abrogates the normal hyperpolarization-induced “sag”
current, suggesting it is the primary HCN in these cells. Future experiments will include using
CRISPR-Cas9 to “knock-out” HCN genes in
Helobdella
and determine the effect on
mechanosensation, as well as using
Hirudo
ex vivo ganglion preparations and primary cell
culture to determine how the loss of HCN activity changes their response to mechanical stimuli.