Magneto is ineffective in controlling electrical properties of cerebellar Purkinje cells

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Abstract

It was recently reported that a magnetic actuator, Magneto, can control neuronal firings at magnetic strength as low as 50 mT (ref. 1), offering an exciting non-invasive approach to manipulating neuronal activity in a variety of research and clinical applications. We investigated whether Magneto can be used to manipulate electric properties of Purkinje cells in the cerebellum, which play critical roles in motor learning and emotional behaviors2. Surprisingly, we found that the application of a magnetic field did not change any electrical properties of Purkinje cells expressing Magneto, raising serious doubt about the previous claim that Magneto can readily be used as a magnetic actuator1.

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Fig. 1: Magnet does not alter membrane potential in Magneto-expressing PCs.

Data availability

The data that support the findings of this study are available from the corresponding author upon reasonable request.

Code availability

No custom algorithm, software, or code was used in the present work.

References

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Acknowledgments

We thank Z.-Y. Feng for her suggestions in statistics and the Core Facilities of Zhejiang University Institute of Neuroscience for technical assistance. This work was supported by grants from the National Natural Science Foundation of China (grant nos. 81625006 and 31820103005) to Y.S. and (grant nos. 31771197 and 91632303) to F.Q.X., the Natural Science Foundation of Zhejiang Province (grant no. Z15C090001) to Y.S. and (grant no. LQ17C090001) to N.W., the Non-profit Central Research Institute Fund of Chinese Academy of Medical Sciences (grant nos. 2017PT31038 and 2018PT31041) to Y.S. and China Postdoctoral Science Foundation (grant no. 2018M640550) to L.Z.

Author information

F.X.X., L.Z. and Y.S. designed the experiments. F.X.X., L.Z., X.T.W., F.J., K.Y.M., N.W. and F.Q.X. collected the data. L.L. supplied reagents. F.X.X., L.Z., X.T.W., K.Y.M. and Y.S. analyzed the data. F.X.X., L.Z., X.T.W., F.J., L.L. and Y.S. wrote the paper.

Correspondence to Ying Shen.

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The authors declare no competing interests.

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Integrated supplementary information

Supplementary Figure 1 The measurement of magnetic intensity.

The inset picture shows that a N52 neodymium 1/16" × 1/4" cylinder magnet was connected to a 1.0-mm diameter (O.D.) borosilicate glass through a white heat shrinkable tube. The plot shows magnetic intensities against the distances of magnets in aCSF (n = 6 measurements). Data are presented as mean ± SEM.

Supplementary Figure 2 Magnet does not change Rinput and capacitance of Magneto-expressing PCs.

(a) Rinput values of individual control PCs (n = 14 cells from 5 animals), the same cells present in Fig. 1e, at the minimal and the maximal magnet strengths. Rinput values at max and min2 stages were normalized to corresponding values at min1 stage. max: 107.0 ± 5.2%. min2: 103.9 ± 3.4%. Values of one-way ANOVA test were F(2, 39) = 0.48, 0.62 (min1 vs max), F(2, 39) = 0.97, 0.39 (min2 vs max), and F(2, 39) = 0.97, 0.39 (min1 vs min2). (b) Rinput of individual Magneto-expressing PCs (n = 15 cells from 6 animals), the same cells present in Fig. 1f, with the minimal and maximal magnet strengths. Likewise, data points were normalized to the corresponding value at min1 stage and percentage changes are shown. max: 101.9 ± 2.4%. min2: 102.6 ± 2.5%. Values of one-way ANOVA test were F(2, 42) = 0.50, 0.61 (min1 vs max), F(2, 42) = 0.26, 0.77 (min2 vs max), and F(2, 42) = 0.26, 0.77 (min1 vs min2). (c) Capacitance of individual control PCs (n = 14 cells from 5 animals), the same cells present in Fig. 1e, at the minimal and maximal magnetic strengths. Percentage changes of capacitance at max and min2 stages were normalized to the corresponding value at min1 stage. max: 100.0 ± 2.2%. min2: 102.2 ± 2.5%. Values of one-way ANOVA test were F(2, 39) = 0.32, 0.73 (min1 vs max), F(2, 39) = 0.005, 1.0 (min2 vs max), and F(2, 39) = 0.005, 1.0 (min1 vs min2). (d) Capacitance of individual Magneto-expressing PCs, the same cells in Fig. 1f, with the minimal and maximal magnet strengths (n = 15 cells from 6 animals). Percentage changes of capacitance at max and min2 stages were normalized to the corresponding value at min1 stage. max: 102.9 ± 2.2%. min2: 106.3 ± 3.4%. Values of one-way ANOVA test were F(2, 42) = 2.1, 0.14 (min1 vs max), F(2, 42) = 0.45, 0.64 (min2 vs max), and F(2, 42) = 0.45, 0.64 (min1 vs min2). Data are presented as mean ± SEM. n.s., not significant.

Supplementary Figure 3 Magnet does not alter spontaneous firings in control PCs.

(a) A schematic illustration of cell-attached recording in PCs. (b) Representative traces of spontaneous firings in control PCs with the minimal and maximal magnetic strengths. The dash lines represent spontaneous events. (c) Firing rates of individual control PCs with the minimal and maximal magnetic strengths. The mean value at the maximal stage was 98.0 ± 1.5% [n = 14 cells from 4 animals; two-sided paired t test; t(13) = 0.01, 1.0]. (d) CV values of individual control PCs at two stages of magnetic strengths. The mean value at the maximal stage was 99.6 ± 2.3% [n = 14 cells from 4 animals; two-sided paired t test; t(13) = 0.17, 0.87]. (e) CV2 values of individual control PCs at two stages of magnetic strength. The mean value at the maximal stage was 102.4 ± 1.8% [n = 14 cells from 4 animals; two-sided paired t test; t(13) = 1.3, 0.22]. (f) LvR values of individual control PCs at two stages of magnetic strength. The mean value at the maximal stage was 99.0 ± 0.5% [n = 14 cells from 4 animals; two-sided paired t test; t(13) = 2.0, 0.07]. Data are presented as mean ± SEM. n.s., not significant.

Supplementary Figure 4 Magnet does not alter spontaneous firings in Magneto-expressing PCs.

(a) Representative traces of spontaneous firings in PCs expressing Magneto with the minimal and maximal magnetic strengths. The dash lines below raw traces represent spontaneous events, indicating that the regularity of the events was not changed by the magnet. (b) Firing rates of individual Magneto-expressing PCs at the minimal and maximal stages of magnetic strength. The mean value of percentage changes at the maximal stage was 101.9 ± 1.6% of that at the minimal stage [n = 10 cells from 4 animals; two-sided paired t test; t(9) = 1.2, 0.26]. (c) CV values of individual Magneto-expressing PCs at the minimal and maximal stages of magnetic strength. The mean value at the maximal stage was 102.0 ± 2.0% [n = 10 cells from 4 animals; two-sided paired t test; t(9) = 1.0, 0.32]. (d) CV2 values of individual Magneto-expressing PCs at two stages. The mean value at the maximal stage was 99.9 ± 2.9% of that at the minimal stage [n = 10 cells from 4 animals; two-sided paired t test; t(9) = 0.03, 0.98]. (e) LvR values of individual Magneto-expressing PCs at two stages of magnetic strength. The mean value at the maximal stage was 99.5 ± 1.7% of that at the minimal stage [n = 10 cells from 4 animals; two-sided paired t test; t(9) = 0.3, 0.77]. Data are presented as mean ± SEM. n.s., not significant.

Supplementary Figure 5 Magnet does not alter action potential initiation in control PCs.

(a) Representative APs obtained by injecting a depolarizing current (50 ms; range 0.1–0.3 nA)10 in control PCs with the minimal and maximal magnetic strengths. The cyan arrowheads show the measurement of threshold, amplitude, FWHM, and AHP. (b) Thresholds of individual control PCs with the minimal and maximal magnetic strengths. The mean value at the maximal stage was 98.4 ± 0.7% [n = 12 cells from 5 animals; two-sided paired t test; t(11) = 1.6, 0.13]. The right panel shows the phase plane plot for single action potential from the same cell with the minimal (grey) or maximal (black) magnetic strengths. (c) AP parameters of individual control PCs at the minimal and maximal stages of magnetic strength. The mean values at the maximal stage were: AP amplitude, 101.2 ± 1.1% [two-sided paired t test; t(11) = 1.1, 0.28]; FWHM: 101.2 ± 2.9 mV [two-sided paired t test; t(11) = 0.5, 0.67]; AHP: 102.7 ± 2.7% [two-sided paired t test; t(11) = 1.0, 0.33]. (d) Sample population spikes of one control PC with the minimal and maximal magnetic stimulation in response to a 300-ms current injection (200 pA). The bar graph indicates ISI values of individual control PCs with the minimal and maximal magnetic strengths. The mean value at the maximal stage was 100.7 ± 1.9% [n = 10 cells from 4 animals; two-sided paired t test; t(9) = 0.38, 0.71]. (e) Input-output curves for control PCs (n = 7 cells from 4 animals) showing numbers of action potentials as a function of injected currents with a duration of 300 ms. The equations for two regression lines were y = 0.153x-14.5 (min) and y = 0.150x-12.9 (max) (p = 0.77; covariant correlation test). Data are presented as mean ± SEM. n.s. not significant.

Supplementary Figure 6 Magnet does not alter action potential initiation in Magneto-expressing PCs.

(a) Representative APs obtained by injecting a depolarizing current in Magneto-expressing PCs with the minimal and maximal magnetic strengths. (b) Thresholds of individual Magneto-expressing PCs at the minimal and maximal stages of magnetic strength. The mean value at the maximal stage was 98.9 ± 0.5% [n = 13 cells from 5 animals; two-sided paired t test; t(12) = 2.1, 0.06]. The right panel shows the phase plane plot (Vm vs dV/dt) for action potentials from the same cell with the minimal or maximal magnetic strength, indicating the pattern of whole AP was not changed by the magnetic stimulation. (c) The parameters of AP amplitude, FWHM, and AHP. The mean values at the maximal stage were: amplitude, 104.3 ± 1.2% [two-sided paired t test; t(12) = 1.9, 0.07]; FWHM: 97.2 ± 1.7% [two-sided paired t test; t(12) = 1.7, 0.11]; AHP: 104.7 ± 2.4% [two-sided paired t test t(12) = 2.1, 0.06]. n = 13 from 5 animals for all parameters. (d) Sample population spikes of one Magneto-expressing PC with the minimal and maximal magnetic stimulation in response to a 300-ms current injection. The mean value of percentage changes at the maximal stage was 100.2 ± 1.8% [n = 9 cells from 4 animals; two-sided paired t test; t(8) = 0.12, 0.91]. (e) Input-output curves for Magneto-expressing PCs (n = 8 cells from 4 animals) showing numbers of action potentials as a function of injected currents (300 ms). The equations for two regression lines were y = 0.149x-14.7 (min) and y = 0.147x-14.7 (max) (p = 0.89; covariant correlation test). Data are presented as mean ± SEM. n.s. not significant.

Supplementary Figure 7

Images of uncropped Western blots used for the preparations in Fig. 1.

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