Figure 6

Sensitivity to transmitters of neurons and astrocytes prepared from 3MST-KO, TRPA1-KO, and 3MST/TRPA1-double-KO compared to those from the wild-type rats. a and b. Responses of neurons to N-methyl-d-aspartate (NMDA) and high K⁺ measured by Ca²⁺ imaging. (a) Typical response to 100 μM NMDA and 50 mM KCl (upper panel) and images of cultured neurons prepared from KO and wild-type rats (lower panel). (b) The ratio of responses to NMDA/ high K⁺ stratified by amplitude (F/F₀). c. Kolmogorov–Smirnov cumulative analysis of NMDA/KCl. **p < 0.01, *p < 0.05. d. Student's t-test was used to determine the NMDA/KCl ratio (%) of total cells. WT: 106 ± 0.70, 3MST-KO: 101 ± 0.67, TRPA1-KO: 103 ± 0.86, 3MST/TRPA1-double KO: 100 ± 0.75. **p < 0.01. (e) A typical response of astrocytes to 20 μM Na₂S₃ and 100 μM glutamate (upper panel) and images of cultured astrocytes prepared from KO and wild-type rats (lower panel). (f) percentage of astrocytes responding to Na₂S₃ and glutamate (blue), and glutamate alone (orange). The percentage of cells responded to Na₂S₃ and glutamate in TRPA1-KO and in M/T-KO is significantly different from that in WT cells by the Chi-squared-test, while that of TRPA1-KO is not significantly different from that of M/T-KO. **p < 0.01 and *p < 0.05. Note that cells responded to Na₂S₃ but not to glutamate were observed 9 out of 423 cells in the wild-type, 5 out of 583 in 3MST-KO, 2 out of 210 in TRPA1-KO, and 0 out of 222 in 3MST/TRPA1-double KO. The experiments were repeated at least ten times. The total cells measured were 173–331 (a–d) and 210–583 (f).