Group III mGluR regulation of synaptic transmission at the SC-CA1 synapse is developmentally regulated

Abstract

Group III metabotropic glutamate receptors (mGluRs) reduce synaptic transmission at the Schaffer collateral-CA1 (SC-CA1) synapse in rats by a presynaptic mechanism. Previous studies show that low concentrations of the group III-selective agonist, L-AP4, reduce synaptic transmission in slices from neonatal but not adult rats, whereas high micromolar concentrations reduce transmission in both age groups. L-AP4 activates mGluRs 4 and 8 at much lower concentrations than those required to activate mGluR7, suggesting that the group III mGluR subtype modulating transmission is a high affinity receptor in neonates and a low affinity receptor in adults. The previous lack of subtype selective ligands has made it difficult to test this hypothesis. We have measured fEPSPs in the presence of novel subtype selective agents to address this question. We show that the effects of L-AP4 can be blocked by LY341495 in both neonates and adults, verifying that these effects are mediated by mGluRs. In addition, the selective mGluR8 agonist, DCPG, has a significant effect in slices from neonatal rats but does not reduce synaptic transmission in adult slices. The mGluR4 selective allosteric potentiator, PHCCC, is unable to potentiate the L-AP4-induced effects at either age. Taken together, our data suggest that group III mGluRs regulate transmission at the SC-CA1 synapse throughout development but there is a developmental regulation of the subtypes involved so that both mGluR7 and mGluR8 serve this role in neonates whereas mGluR7 is involved in regulating transmission at this synapse throughout postnatal development.

Figure 1

L-AP4 reduces synaptic transmission at the SC-CA1 synapse in the neonatal rat hippocampus. A: representative field excitatory postsynaptic potentials (fEPSP) recorded from the dendritic layer of CA1 with the use of extracellular field recording techniques. Traces are the average of 5 trials immediately before and 10 min after application of L-AP4 (50µM). Field EPSPs were elicited by stimulating the Schaffer collaterals. B: concentration-response curve for L-AP4. Shown is the depression (mean ± SEM) of fEPSP slope induced by increasing concentrations of L-AP4. Data are expressed as a percentage of predrug fEPSP slope. The concentration producing half-maximal depression is 25µM. N = 3–6 for each point.

Figure 2

High concentrations of L-AP4 are required to reduce synaptic transmission at the SC-CA1 synapse of the adult rat hippocampus. A: representative field excitatory postsynaptic potential (fEPSP) recorded from the dendritic layer of CA1 with the use of extracellular field recording techniques, showing the effects of the group III-selective agonist L-AP4 (50µM or 1mM). Field EPSPs were elicited by stimulating the Schaffer collaterals. Traces are the averages of 5 trials immediately before and 10 min after application of L-AP4. High concentrations of L-AP4 (1mM) are required to reduce the slope of the fEPSP. B: concentration-response curve for L-AP4. Shown is the depression (mean ± SEM) of fEPSP slope induced by increasing concentrations of L-AP4. Data are expressed as a percentage of predrug fEPSP slope. The concentration producing half-maximal depression is 1mM. N = 3–5 for each point.

Figure 3

LY341495 antagonizes the effect of L-AP4 at the SC-CA1 synapse in the neonatal and adult rat hippocampus. A: Representative fEPSPs and bar graph depicting the effect of 10µM L-AP4 on fEPSP slope in the absence and presence of 30µM LY341495 on slices from neonatal rats. B: Representative fEPSPs and bar graph showing the effect of 650µM L-AP4 of fEPSP slope in the absence and presence of 100µM LY341495 on slices from adult rats. Data (mean ± SEM) are expressed as a percentage of predrug value. N = 3–5 for each experiment; * P < 0.05, ** P < 0.01.

Figure 4

DCPG reduces synaptic transmission at the SC-CA1 synapse in the neonatal, but not adult rat hippocampus. A,D: Representative fEPSPs and bar graph depicting the effects of 1µM DCPG on fEPSP slope in hippocampal slices from neonatal rats. C: Concentration response curve for DCPG. Shown is the depression (mean ± SEM) of fEPSP slope induced by increasing concentrations of DCPG. Data are expressed as a percentage of predrug fEPSP slope. The concentration producing half-maximal depression is 688nM. N = 3–7 for each point. B,D: Representative fEPSPs and bar graph depicting the effects of 10µM DCPG in slices from adult rats. Data (mean ± SEM) are expressed as a percentage of predrug value. N = 4–7 for each experiment; * P < 0.05.

Figure 5

PHCCC does not potentiate the effects of L-AP4 at the SC-CA1 synapse in slices from neonatal and adult hippocampus. A: Representative fEPSPs and bar graph depicting the effects of 10µM L-AP4 on fEPSP slope in the presence and absence of 30µM PHCCC on slices from neonatal rats. 10µM L-AP4, 61±3.3% of predrug; 10µM L-AP4 + 30µM PHCCC, 54.5±3.6% of predrug; p>0.05. B: Representative fEPSPs and bar graph depicting the effects of 750µM L-AP4 on fEPSP slope in the presence and absence of 30µM PHCCC on slices from adult rats. 750µM L-AP4, 61.6±7% of predrug; 750µM L-AP5 + 30µM PHCCC, 71±3% of predrug; p>0.05. Data (mean ± SEM) are presented as percentage of predrug value. N = 3–4 for each experiment.

Figure 6

Concentration-response relationship of Z-cyclopentyl AP4 in the presence of PHCCC. Cells lines expressing the group III mGluRs were screened for activation by Z-cyclopentyl AP4. The addition of 10µM PHCCC potentiated the effects of Z-cyclopentyl AP4 at mGluR4 (A, D), but not mGluR8 (B, D). Z-Cyclopentyl AP4 has no effect on mGluR7 (C, D). mGluR4: Z-cyclopentyl AP4 EC₅₀ = 49.13 ± 10.86µM versus Z-cyclopentyl AP4 + PHCCC EC₅₀ = 14.5 ± 5.1 µM, p<0.01. mGluR8: (z)-cyclopentyl AP4 EC₅₀ = 124.9 ± 21 µM versus Z-cyclopentyl AP4 + PHCCC EC₅₀ = 85 ± 9.7 µM, p<0.05. Data (mean ± SEM) are expressed as a percentage of the maximal L-AP4 response for each individual cell line and were calculated based on means of three independent experiments performed in triplicate.

Figure 7

PHCCC does not potentiate the effects of Z-cyclopentyl AP4 at the SC-CA1 synapse in slices from neonatal and adult hippocampus. A (neonatal) and B (adult): Representative fEPSPs, time course and graphic representation showing the effects of Z-cyclopentyl AP4 (50µM) in the presence and absence of PHCCC (30µM). Neonatal: 50µM Z-cyclopentyl AP4 95.6±3.8% of predrug, 50µM Z-cyclopentyl AP4 + 30µM PHCCC 90±4.8% of predrug; p>0.05. Adult: 50µM Z-cyclopentyl AP4 94.5±0.5% of predrug, 50µM Z-cyclopentyl AP4 + 30µM PHCCC 95.4±1% of predrug; p>0.05. Data (mean ± SEM) are presented as percentage of predrug value. N = 3–6 for each experiment.

Figure 8

AMN082 does not reduce synaptic transmission at the SC-CA1 synapse of adult animals. Representative fEPSPs (inset) and time course showing the lack of effect of AMN082 (1µM). Data (mean ± SEM) are presented as percentage of predrug value. N = 5.

Figure 9

AMN082 does not result in activation of GIRK channels in mGluR7/GIRK cells. Increasing concentrations of AMN082 or L-AP4 were added to mGluR7/GIRK cells and thallium flux was measured. L-AP4 EC₅₀ = 43.3 ± 4.1 µM. Data (mean ± SEM) are expressed as a percentage of the maximal L-AP4 response and were calculated based on means of four independent experiments performed in triplicate.