Metabotropic Glutamate 2 Receptors Modulate Synaptic Inputs and Calcium Signals in Striatal Cholinergic Interneurons

LY379268 exerts an inhibitory effect on IPSP recorded from cholinergic interneurons. A, A control IPSP evoked by intrastriatal stimulation in MK-801, CNQX, and BMI (30, 10, and 10 μm, respectively) in the perfusing solution was significantly reduced in amplitude by LY379268 (5 min, 10 μm) in a reversible manner. B, A dose–response curve for the inhibitory effect of LY379268 revealed an IC₅₀ of 1.6 ± 0.5 μm. Each data point was obtained by averaging at least four independent observations. C, The P-type HVA channel blocker ω-Aga-IVA (20 nm) significantly reduced the IPSP amplitude recorded in 30 μm MK-801, 10 μmCNQX, and 10 μm BMI. After reaching a steady-state inhibition with ω-Aga-IVA, coapplication of ω-Aga-IVA plus LY379268 (10 μm) did not induce any further decrease in IPSP amplitude. D, ω-Ctx-GVIA (1 μm) reduced the IPSP amplitude. The following bath application of LY379268 (10 μm) produced an additional decrease in the IPSP amplitude. E, Bath application of oxotremorine (300 nm, 1 min) hyperpolarized the recorded cell and blocked the action potential discharge in a reversible manner. In LY379268 (10 μm, 10 min preincubation), the oxotremorine-mediated effect was not modified.

Glutamate- and GABA-mediated synaptic components are inhibited by group II mGlu receptor activation. A, The glutamate-mediated component of the EPSP was isolated by bathing the slice in a solution containing BMI (10 μm), a GABA_A receptor antagonist, plus scopolamine (1 μm). In this condition, LY379268 (10 μm, 5 min) reduced the EPSP amplitude without affecting intrinsic membrane properties. A complete return to baseline values was observed 10–15 min after washout. B, In the presence of the ionotropic glutamate receptor antagonists MK-801 (30 μm) and CNQX (10 μm) plus scopolamine, the GABAergic component of the synaptic potential was significantly reduced by LY379268 (10 μm, 5 min). This inhibitory effect was fully reversed after 10–15 min washout. Resting membrane potential was constant throughout the experiment (−66 and −60 mV).

Spontaneous calcium-dependent PPs and simultaneous [Ca²⁺]_i transients. A, Intracellular recordings were performed with 1 μm TTX and 10 mm TEA in the perfusing solution and CsCl (2m) plus bis-fura-2 (1 mm) in the recording microelectrode. In this experimental condition, a spontaneous, rhythmic spiking activity was generated (a). Each of these long-lasting PPs was followed by a prominent afterhyperpolarization and was coupled to a simultaneous transient increase in [Ca²⁺]_i, as revealed by combined optical recordings (b). In c, at higher sweep speed (compare time scale inA), a single PP (thick bar) and the coincident [Ca²⁺]_i rise (thin bar) are shown. The dotted line indicates which PP was enlarged in b. B, [Ca²⁺]_i transients were fully blocked by perfusing a cocktail solution; HVA calcium channel blockers were composed of 10 μm nifedipine for L-type, 20 nm ω-Aga-IVA for P-type, 1 μm ω-conotoxin GVIA for N-type, and 1 μm ω-conotoxin MVIIC for Q-type.

LY379268 reduces calcium-dependent PPs by modulating P-type calcium channels. Aa, Spontaneous PPs recorded from a cholinergic interneuron (top) and simultaneous [Ca²⁺]_i elevation (bottom) in control condition and in the presence of LY379268 (10 μm). At higher sweep speed, thetrace represents a single PP in controls and in LY379268 (b). Note the net reduction caused by LY379268 both in PPs duration and increase in [Ca²⁺]_i (b).Ba, Similarly, bath-applied ω-Aga-IVA (20 nm) mimicked the inhibitory effect induced by the mGlu receptor agonist, on both the duration of PP and the concomitant [Ca²⁺]_i rise. The traceon the right (b) shows, at higher sweep speed, the action of ω-Aga-IVA on a single PP and on a individual [Ca²⁺]_i transient.

Additive effects of ω-Aga-IVA and ω-Ctx-GVIA on PPs. ω-Aga-IVA (20 nm, 10 min) reversibly reduced the duration of PPs compared with controls (a,b), as well as the peak amplitude of coincident [Ca²⁺]_i transients. In ω-Aga-IVA, ω-Ctx-GVIA (1 μm) produced an additional decrease in PPs duration, as well as in the simultaneous [Ca²⁺]_i elevation (c).

mGlu2 receptor activation reduces ACh release through the suppression of P-type calcium channels. A, A dose–response curve for the inhibitory effect of the group II mGlu agonist DCG-IV on the electrically evoked release of endogenous ACh from a striatal slice preparation, expressed as the St₂/St₁ ratio values (electrical stimulation after and before agonist administration, respectively). Each data point was obtained from at least six independent observations. All experiments were performed in the presence of 30 μm MK-801. B, Summary plots showing that the inhibitory action of LY379268 was mimicked and occluded by ω-Aga-IVA. In slices preincubated in ω-Aga-IVA, LY379268 (ratio St2/St1) was no more effective in the modulation of ACh release compared with controls.