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Developmental waves of mechanosensitivity acquisition in sensory neuron subtypes during embryonic development.

Lechner SG, Frenzel H, Wang R, Lewin GR - EMBO J. (2009)

Bottom Line: Sensory neurons that are mechanoreceptors or proprioceptors acquire mature mechanotransduction indistinguishable from the adult already at E13.In contrast, most nociceptive (pain sensing) sensory neurons acquire mechanosensitive competence as a result of exposure to target-derived nerve growth factor.The highly regulated process of mechanosensory acquisition unveiled here, reveals new strategies to identify molecules required for sensory neuron mechanotransduction.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience, Max-Delbrück Center for Molecular Medicine, Berlin-Buch, Germany.

ABSTRACT
Somatic sensation relies on the transduction of physical stimuli into electrical signals by sensory neurons of the dorsal root ganglia. Little is known about how and when during development different types of sensory neurons acquire transduction competence. We directly investigated the emergence of electrical excitability and mechanosensitivity of embryonic and postnatal mouse sensory neurons. We show that sensory neurons acquire mechanotransduction competence coincident with peripheral target innervation. Mechanotransduction competence arises in different sensory lineages in waves, coordinated by distinct developmental mechanisms. Sensory neurons that are mechanoreceptors or proprioceptors acquire mature mechanotransduction indistinguishable from the adult already at E13. This process is independent of neurotrophin-3 and may be driven by a genetic program. In contrast, most nociceptive (pain sensing) sensory neurons acquire mechanosensitive competence as a result of exposure to target-derived nerve growth factor. The highly regulated process of mechanosensory acquisition unveiled here, reveals new strategies to identify molecules required for sensory neuron mechanotransduction.

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Sequential acquisition of mechanosensitivity in sensory neurons. (A) Example traces of mechanosensitive currents recorded from embryonic sensory neurons. Mechanical stimuli were applied (blue trace), and based on their inactivation time constant currents were classified as RA-, IA- and SA-currents. (B) Stacked histogram showing proportions of sensory neurons with mechanically activated currents at different developmental stages. Significant changes in proportions occur between E12.5 and E13.5 and between E18 and P0, respectively (***P<0.001; *P<0.05, χ2 test). (C) Phase-contrast photomicrographs showing acutely dissociated E12.5 and E13.5 cultures, large neurons are only present at E13.5 (red arrows). (D) RA-current proportions are plotted separately for large neurons (>14 μm, dark blue) and small neurons (<14 μm, pale blue). (E) Single-cell real-time PCR from neurons with an RA-current. Functionality of primers was confirmed using plasmids containing trk-receptor cDNAs and cDNA from whole E13.5 DRGs (Supplementary Figure S3).
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f2: Sequential acquisition of mechanosensitivity in sensory neurons. (A) Example traces of mechanosensitive currents recorded from embryonic sensory neurons. Mechanical stimuli were applied (blue trace), and based on their inactivation time constant currents were classified as RA-, IA- and SA-currents. (B) Stacked histogram showing proportions of sensory neurons with mechanically activated currents at different developmental stages. Significant changes in proportions occur between E12.5 and E13.5 and between E18 and P0, respectively (***P<0.001; *P<0.05, χ2 test). (C) Phase-contrast photomicrographs showing acutely dissociated E12.5 and E13.5 cultures, large neurons are only present at E13.5 (red arrows). (D) RA-current proportions are plotted separately for large neurons (>14 μm, dark blue) and small neurons (<14 μm, pale blue). (E) Single-cell real-time PCR from neurons with an RA-current. Functionality of primers was confirmed using plasmids containing trk-receptor cDNAs and cDNA from whole E13.5 DRGs (Supplementary Figure S3).

Mentions: There are three types of mechanically activated currents in adult mouse DRG neurons. These currents can be readily distinguished by their inactivation kinetics (Figure 2A) and are classified into RA-type, IA-type and SA-type currents (McCarter et al, 1999; Drew et al, 2002; Hu and Lewin, 2006; McCarter and Levine, 2006). We recorded mechanosensitive currents by stimulating the soma with very small (⩾350 nm) and rapidly applied (3.5 μm/ms) displacement stimuli. At E11.5 and E12.5, none of the cells tested responded with inward current to mechanical stimulation (Figure 2B). Just 1 day later, at E13.5, 59% (42/71 tested neurons) exhibited mechanically activated currents (Figure 2B). Interestingly, at this stage, mechanosensitivity was largely restricted to a subpopulation of large diameter neurons (Figure 2C and D), which had also just appeared at E13.5 (Figure 2C, for size frequency plots see Supplementary Figure S2A); neurons smaller than 14 μm did not possess a mechanosensitive current at this stage (Figure 2D). Almost all neurons with a mechanosensitive current exhibited RA-currents (88.1%), whereas IA- and SA-currents were found in only 7.2 and 4.7% of the recorded cells, respectively (Figure 2B). The incidence and distribution of mechanosensitive currents at E14.5 was virtually the same as at E13.5. Many of the neurons recorded at E13.5 possessed a humped AP, but the AP width was still narrow compared with more mature nociceptors (Figure 1B and C). We, thus, carried out single-cell PCR experiments to characterize neurotrophin receptor expression in functionally characterized neurons with an RA-type current at E13.5. Of 15 large diameter neurons recorded with a mechanosensitive current (mean cell diameter 17.6±0.9 μm), all expressed either TrkC (5 cells), TrkB (5 cells) or both (5 cells) (Figure 2E). Importantly, TrkA transcripts could not be detected in any of the 15 mechanosensitive neurons tested. The control experiments demonstrated that the primers used to detect TrkA transcripts were equally efficient in amplifying TrkA as the TrkB and TrkC primers used (Supplementary Figure S3). Thus, the first neurons to acquire mechanosensitivity have the molecular make-up predicted for low threshold mechanoreceptors (Figure 2E).


Developmental waves of mechanosensitivity acquisition in sensory neuron subtypes during embryonic development.

Lechner SG, Frenzel H, Wang R, Lewin GR - EMBO J. (2009)

Sequential acquisition of mechanosensitivity in sensory neurons. (A) Example traces of mechanosensitive currents recorded from embryonic sensory neurons. Mechanical stimuli were applied (blue trace), and based on their inactivation time constant currents were classified as RA-, IA- and SA-currents. (B) Stacked histogram showing proportions of sensory neurons with mechanically activated currents at different developmental stages. Significant changes in proportions occur between E12.5 and E13.5 and between E18 and P0, respectively (***P<0.001; *P<0.05, χ2 test). (C) Phase-contrast photomicrographs showing acutely dissociated E12.5 and E13.5 cultures, large neurons are only present at E13.5 (red arrows). (D) RA-current proportions are plotted separately for large neurons (>14 μm, dark blue) and small neurons (<14 μm, pale blue). (E) Single-cell real-time PCR from neurons with an RA-current. Functionality of primers was confirmed using plasmids containing trk-receptor cDNAs and cDNA from whole E13.5 DRGs (Supplementary Figure S3).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2664657&req=5

f2: Sequential acquisition of mechanosensitivity in sensory neurons. (A) Example traces of mechanosensitive currents recorded from embryonic sensory neurons. Mechanical stimuli were applied (blue trace), and based on their inactivation time constant currents were classified as RA-, IA- and SA-currents. (B) Stacked histogram showing proportions of sensory neurons with mechanically activated currents at different developmental stages. Significant changes in proportions occur between E12.5 and E13.5 and between E18 and P0, respectively (***P<0.001; *P<0.05, χ2 test). (C) Phase-contrast photomicrographs showing acutely dissociated E12.5 and E13.5 cultures, large neurons are only present at E13.5 (red arrows). (D) RA-current proportions are plotted separately for large neurons (>14 μm, dark blue) and small neurons (<14 μm, pale blue). (E) Single-cell real-time PCR from neurons with an RA-current. Functionality of primers was confirmed using plasmids containing trk-receptor cDNAs and cDNA from whole E13.5 DRGs (Supplementary Figure S3).
Mentions: There are three types of mechanically activated currents in adult mouse DRG neurons. These currents can be readily distinguished by their inactivation kinetics (Figure 2A) and are classified into RA-type, IA-type and SA-type currents (McCarter et al, 1999; Drew et al, 2002; Hu and Lewin, 2006; McCarter and Levine, 2006). We recorded mechanosensitive currents by stimulating the soma with very small (⩾350 nm) and rapidly applied (3.5 μm/ms) displacement stimuli. At E11.5 and E12.5, none of the cells tested responded with inward current to mechanical stimulation (Figure 2B). Just 1 day later, at E13.5, 59% (42/71 tested neurons) exhibited mechanically activated currents (Figure 2B). Interestingly, at this stage, mechanosensitivity was largely restricted to a subpopulation of large diameter neurons (Figure 2C and D), which had also just appeared at E13.5 (Figure 2C, for size frequency plots see Supplementary Figure S2A); neurons smaller than 14 μm did not possess a mechanosensitive current at this stage (Figure 2D). Almost all neurons with a mechanosensitive current exhibited RA-currents (88.1%), whereas IA- and SA-currents were found in only 7.2 and 4.7% of the recorded cells, respectively (Figure 2B). The incidence and distribution of mechanosensitive currents at E14.5 was virtually the same as at E13.5. Many of the neurons recorded at E13.5 possessed a humped AP, but the AP width was still narrow compared with more mature nociceptors (Figure 1B and C). We, thus, carried out single-cell PCR experiments to characterize neurotrophin receptor expression in functionally characterized neurons with an RA-type current at E13.5. Of 15 large diameter neurons recorded with a mechanosensitive current (mean cell diameter 17.6±0.9 μm), all expressed either TrkC (5 cells), TrkB (5 cells) or both (5 cells) (Figure 2E). Importantly, TrkA transcripts could not be detected in any of the 15 mechanosensitive neurons tested. The control experiments demonstrated that the primers used to detect TrkA transcripts were equally efficient in amplifying TrkA as the TrkB and TrkC primers used (Supplementary Figure S3). Thus, the first neurons to acquire mechanosensitivity have the molecular make-up predicted for low threshold mechanoreceptors (Figure 2E).

Bottom Line: Sensory neurons that are mechanoreceptors or proprioceptors acquire mature mechanotransduction indistinguishable from the adult already at E13.In contrast, most nociceptive (pain sensing) sensory neurons acquire mechanosensitive competence as a result of exposure to target-derived nerve growth factor.The highly regulated process of mechanosensory acquisition unveiled here, reveals new strategies to identify molecules required for sensory neuron mechanotransduction.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience, Max-Delbrück Center for Molecular Medicine, Berlin-Buch, Germany.

ABSTRACT
Somatic sensation relies on the transduction of physical stimuli into electrical signals by sensory neurons of the dorsal root ganglia. Little is known about how and when during development different types of sensory neurons acquire transduction competence. We directly investigated the emergence of electrical excitability and mechanosensitivity of embryonic and postnatal mouse sensory neurons. We show that sensory neurons acquire mechanotransduction competence coincident with peripheral target innervation. Mechanotransduction competence arises in different sensory lineages in waves, coordinated by distinct developmental mechanisms. Sensory neurons that are mechanoreceptors or proprioceptors acquire mature mechanotransduction indistinguishable from the adult already at E13. This process is independent of neurotrophin-3 and may be driven by a genetic program. In contrast, most nociceptive (pain sensing) sensory neurons acquire mechanosensitive competence as a result of exposure to target-derived nerve growth factor. The highly regulated process of mechanosensory acquisition unveiled here, reveals new strategies to identify molecules required for sensory neuron mechanotransduction.

Show MeSH
Related in: MedlinePlus