Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2013 Dec 10;110(50):20302-7.
doi: 10.1073/pnas.1311686110. Epub 2013 Nov 25.

T-type Channel Blockade Impairs Long-Term Potentiation at the Parallel fiber-Purkinje Cell Synapse and Cerebellar Learning

Affiliations
Free PMC article

T-type Channel Blockade Impairs Long-Term Potentiation at the Parallel fiber-Purkinje Cell Synapse and Cerebellar Learning

Romain Ly et al. Proc Natl Acad Sci U S A. .
Free PMC article

Abstract

CaV3.1 T-type channels are abundant at the cerebellar synapse between parallel fibers and Purkinje cells where they contribute to synaptic depolarization. So far, no specific physiological function has been attributed to these channels neither as charge carriers nor more specifically as Ca(2+) carriers. Here we analyze their incidence on synaptic plasticity, motor behavior, and cerebellar motor learning, comparing WT animals and mice where T-type channel function has been abolished either by gene deletion or by acute pharmacological blockade. At the cellular level, we show that CaV3.1 channels are required for long-term potentiation at parallel fiber-Purkinje cell synapses. Moreover, basal simple spike discharge of the Purkinje cell in KO mice is modified. Acute or chronic T-type current blockade results in impaired motor performance in particular when a good body balance is required. Because motor behavior integrates reflexes and past memories of learned behavior, this suggests impaired learning. Indeed, subjecting the KO mice to a vestibulo-ocular reflex phase reversal test reveals impaired cerebellum-dependent motor learning. These data identify a role of low-voltage activated calcium channels in synaptic plasticity and establish a role for CaV3.1 channels in cerebellar learning.

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.
Activation of T-type calcium channels is necessary to induce PF–PC LTP but not for LTD. (A and B) T-type calcium channels are required for LTP induction. (A, Upper) Representative traces from 10 successive sweeps before and 30 min after induction of LTP. (A, Lower) Time course of normalized EPSC charge in control (black), in the continuous presence of 1 µM TTA-P2 (blue), and in CaV3.1−/− mice (red). The LTP induction protocol started at time 0. (B) EPSC charge before and 30 min after LTP induction. Filled symbols represent individual cells, whereas empty symbols represent means. Black, red, and blue indicate results from control mice, from CaV3.1−/− mice, and in the presence of TTA-P2, respectively, and corresponding n = 8, 7, and 7 individual experiments for the three sets of mice, each run on a different slice. Note that data from some cells are superimposed. (C and D) T-type calcium channels are not required for LTD induction. (C, Upper) Representative traces from 10 successive sweeps before and 30 min after induction of LTD. (C, Lower) Time course of normalized EPSC charge in control (black), 1 µM TTA-P2 (blue), and in CaV3.1−/− mice (red). The LTD induction protocol started at time 0. (D) EPSC charge before and 30 min after LTD induction. n = 7 individual experiments from different slices for the three sets of conditions.
Fig. 2.
Fig. 2.
Effects of T-type calcium channel functional inactivation on the spontaneous activity of PCs. (A–C) T-type calcium channel blockade slightly reduces the firing activity of PCs in vitro. (A) Representative traces from extracellular recordings of a PC before and after application of TTA-P2 in slices. (B) Normalized mean firing frequency and (C) coefficient of variation of the ISI in control and in TTA-P2 conditions. Paired experiments run on 16 cells and 13 cells, respectively. *P < 0.05. (D) PC firing is altered in vivo. Cumulative distribution function and boxplot of the PCs’ firing frequencies and CVISI (Left and Right, respectively). Boxplots represent minimum, first quartile, median, third quartile, and maximum values of the distribution. Filled circles represent outliers; the mean value is represented by a plus in the boxplot. Tests run on 99 and 144 cells recorded in control and CaV3.1 KO mice, respectively; statistical P values, *P < 0.05, **P < 0.01.
Fig. 3.
Fig. 3.
Motor behavioral deficit after inactivation of T-type channels. Coordination was significantly impaired in CaV3.1−/− mice (A–C) comparison of motor performance of WT and CaV3.1−/− mice (black and red symbols, respectively, and n = 16 and 12). For details on the individual tests, see Methods. (A) Motor impairment in the elevated beam test. Mice were made to walk on a horizontal beam. Time to perform on a 100-cm-long run was measured as well as the number of times hindlimbs slipped off the beam during three trials. CaV3.1−/− mice take more time to achieve the test (P < 0.01, repeated measures ANOVA) (A, Left) and stumbled more often than WT animals (the rear leg slips off the bar) (P < 0.01, repeated measures ANOVA) (A, Right)). Mice performance increased during the three trials for both the run time and the number of slips, F(2, 52) > 20, P < 0.001, for both phenotypes. (B) Motor impairment in the pole test. The animal was positioned with head up close to the pole top. It had to turn before going head down the rod to return to its cage. TT measured the delay until the mouse was head down. Mutant animals took longer time to reach the box (TT = 39 ± 11 s for CaV3.1−/− and 22 ± 7 s for WT, P < 0.001) (B, Left). There is a slight but nonsignificant increase of CaV3.1−/− mice that could not achieve the task (B, Right). (C) Motor impairment in the rotarod test. The time CaV3.1−/− mice stayed on the rod with a constant rotation speed (10 rpm or 20 rpm) or with accelerating speed (from 4 to 40 rpm in 2 min) was significantly shorter for CaV3.1−/− (10 rpm, P < 0 0.001; 20 rpm, P < 0 0.001; accelerated protocol, P < 0.001; repeated measures ANOVA).
Fig. 4.
Fig. 4.
Cerebellar learning is affected by manipulation of CaV3.1 function. Short- (A) and long-term adaptation (B) of the VOR gain was induced by mismatched vestibular and visual stimulation. (A) Five 10-min sessions of in-phase drum and table stimulation resulted in a decrease of VOR gain to 56% ± 4% of baseline value (n = 8). A similar decrease was seen in CaV3.1 KO mice (59% ± 3%, n = 12, P = 0.95 for the curve, repeated measures ANOVA), but not in mice injected with TTA-P2 (67% ± 7%, n = 7, P = 0.010 for the curve, repeated measures ANOVA). Consolidation of VOR gain decrease was found not to be different between groups (P = 0.75, one-way ANOVA). (B) Next, mice were subjected to 3 additional days with increasing amplitude of visual stimulation during the training sessions, aimed at reversing VOR phase. This highly demanding task revealed significant deficits in cerebellar learning in CaV3.1 KO mice (days 3 and 4, P < 0.002, repeated measures ANOVA) and in mice injected with TTA-P2 (day 2, P = 0.044; days 3 and 4, P < 0.0001, repeated measures ANOVA).

Similar articles

See all similar articles

Cited by 30 articles

See all "Cited by" articles

Publication types

Substances

Feedback