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Abstract

The retrosplenial cortex (RSC) plays an important role in navigation, memory and pain. However, there are few studies on excitatory synaptic transmission in the RSC. Here, we used a multi-electrode array recording system (MED64) to study the characteristics of excitatory synaptic transmission in the RSC and the contribution of different types of voltage-gated Ca²⁺ channels (VGCCs) in excitatory synaptic transmission. We found that glutamate is the major excitatory transmitter for RSC, and postsynaptic alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors contribute to excitatory synaptic transmission. We also found that the N-type calcium channel blocker ω-conotoxin GVIA (ω-Ctx GVIA) had an inhibitory effect on basal synaptic transmission. The inhibitory effect was not consistent across channels, suggesting the actions effect of N-type VGCCs in RSC was inhomogeneous in spatial distribution. Our findings provide strong evidence that excitatory synaptic transmission in the RSC is mainly mediated by AMPA receptors and that N-type VGCCs mediate fast synaptic transmission in the RSC of adult mice.

Keywords

Retrosplenial cortex synaptic transmission AMPA receptor voltage-gated calcium channels ω-conotoxin GVIA

Introduction

It is known that the retrosplenial cortex (RSC) is one of the core areas of the cerebral cortex that supports higher cognitive functions and is involved in sensorimotor and cognitive functions.^1–5 For example, it has been demonstrated that the RSC is involved in nociceptive transmission and modulation.⁶ Human and animal imaging studies found that migraine and neuropathic pain may lead to enhanced functional connectivity of the RSC with the anterior cingulate cortex (ACC)^7,8 a key cortical area for pain perception.^9,10 In consistent with this notion, our recent study has found that there is direct excitatory synaptic connection between the RSC and the ACC, and activation of this connection optogenetically facilitated behavioral nociceptive responses.¹¹ However, there are few electrophysiological studies on excitatory synaptic transmission in the RSC.

Glutamate receptors are necessary for normal synaptic transmission.^12,13 Accumulative studies have consistently shown that glutamatergic synapses not only play an important role in sensory transmission, including pain and itch transmission, but also contribute to different levels of injury sensitization in the brain.¹⁴ There are three main subtypes of ionotropic glutamate receptors: alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor, N-methyl-D-aspartate (NMDA) receptor and Kainate (KA) receptor.¹³ A number of studies have shown that AMPA and KA receptors are involved in mediating rapid postsynaptic responses in the ACC.^9,15,16 Patch-clamp recordings from genetically altered mice indicate that postsynaptic KA receptors play a role in rapid synaptic transmission within pyramidal neurons of the ACC.¹⁷ In addition, a recent study has shown that calcium-permeably AMPA receptors contributes to the potentiation of synaptic plasticity.¹⁸ However, the detailed roles of these three types of glutamate receptors in the excitatory synaptic transmission of the RSC are still unclear.

Voltage-gated Ca²⁺ channels (VGCCs) have a central role in neuronal synaptic transmission. Previous studies have identified various types of calcium channels in peripheral and central nervous system.¹⁹ However, few studies have been reported on the synaptic transmission of VGCCs in the RSC. Thus, we used a 64-channel multi-electrode dish (MED64) system to characterize the roles of different glutamate receptors as well as VGCCs in basal excitatory synaptic transmission of the RSC in adult mice. The MED64 system allows us to simultaneously detect fEPSPs at multiple sites in the mouse RSC, which is difficult with conventional field recording systems.^20–22 We found that excitatory synaptic transmission within the RSC is mediated by glutamate and its receptors, with AMPA receptors making a major contribution. We also found that N-type VGCCs play a dominant role in RSC synaptic transmission.

Methods

Animals

Adult male C57BL/6 mice (6–8 weeks old) were purchased from the Beijing Vital River Laboratory Animal Technology Co., Ltd. All mice were randomly housed in corncob-lined plastic cages under an artificial 12 h light/12 h dark cycle (lights on 9 a.m. to 9 p.m.) with enough food and water. Animal protocols were approved by the Ethics Committee of Oujiang Laboratory.

Brain slice preparation

Mice were anesthetized with 1%–2% isoflurane and sacrificed by decapitation. Coronal brain slices (300 μm) containing the RSC from C57BL/6 mice were prepared at 4°C using standard methods.²⁰ The whole brain was removed from the skull and submerged in the ice-cold oxygenated (95% O₂ and 5% CO₂) artificial cerebrospinal fluid (ACSF) containing (in mM) 124 NaCl, 2.5 KCl, 2 CaCl₂, 2 MgSO₄, 25 NaHCO₃, 1 NaH₂PO₄, and 10 glucose, pH 7.3–7.4. After cooling for a short moment, the whole brain was then cut into appropriate sections to be placed on the tissue slicer of the Leica VT1200S vibratome on the cutting stage. Slices were transferred to a submerged recovery chamber with oxygenated (95% O₂ and 5% CO₂) ACSF at room temperature for at least 1 h.

Preparation of the multielectrode array

A commercial MED64 recording system (Panasonic) was used for extracellular field potential recordings. The procedure for preparation of the MED64 probe (P515A, Panasonic) used standard methods.²⁰ The MED64 probe has an array of 64 planar microelectrodes, each arranged in an 8 × 8 pattern, with an interelectrode distance of 150 mm. Before use, the surface of the MED64 probe was treated with 0.1% polyethyleneimine (P-3143, Sigma-Aldrich) in 25 mmol/L borate buffer, pH 8.4, overnight at room temperature. The surface of the probe has been rinsed with sterile distilled water at least three times before using the probe in the experiments.

Field potential recording in adult RSC slices

After incubation, one slice containing the RSC was transferred to the prepared MED64 probe and perfused with the oxygenated fresh ACSF at room temperature and maintained at a flow rate of 2 mL/min. The slice was positioned on the MED64 probe in such a way that the different layers of the RSC were entirely covered by the whole array of the electrodes, and then a fine-mesh anchor was placed on the slice to ensure its stabilization during the experiments. After at least a 1 h recovery period for the slices in the recording chamber, biphasic constant-current pulse stimulation (0.2 ms) was applied to the stimulation channel. The stimulation intensity was adjusted according to the half-maximal field excitatory postsynaptic potential (fEPSPs) was elicited in the channels closest to the stimulation site. The parameter of ‘slope’ indicated the average slope of each fEPSPs recorded by activated channels. Stable baseline responses were first recorded until the baseline response variation was less than 5% in most of the active channels within 30 min.

Data analysis

The data is presented as means ± standard error of the mean (SEM). Statistical comparisons between two groups were performed using two-tail paired Student’s t-test, one-way analysis of variance (ANOVA) to identify significant differences. In all cases, p < 0.05 was considered statistically significant.

Drug

Drugs were freshly prepared: The selective competitive NMDA receptor antagonist D-AP5, the non-competitive AMPA receptor antagonist GYKI 53655 hydrochloride and the selective non-NMDA ionotropic glutamate receptor antagonist CNQX were purchased from Tocris Cookson (Bristol, UK). N-type VGCCs blocker ω-Ctx GVIA, P/Q-type VGCCs blocker ω-Agatoxin IVA (ω-Aga IVA), L-type VGCCs blocker nimodipine and R-type VGCCs blocker SNX-482 were purchased from MedChemExpress (New Jersey, USA). T-type VGCCs blocker NiCl₂, all types of VGCCs blocker CdCl₂ were obtained from Sigma. Drugs were stored in frozen aliquots at −20°C. All drugs were diluted from the stock solutions to the final desired concentration in the ACSF before immediate use.

Results

Spatial distribution of extracellular fEPSPs in the RSC

In the present study, a 64-channel multielectrode array known as MED64 was used to record the spatial distribution of extracellular fEPSPs within RSC of adult mice.^20,21,23,24 An RSC slice was placed on top of the 8 × 8 square-shaped MED64 probe electrodes (Figure 1(a)). A channel in the RSC was selected to stimulate the slice (Figure 1(b), red circle). The remaining 63 channels recorded fEPSPs with intensity dependence. This can be seen in a single channel and the remaining channels (Figure 1(c)–(e)). The majority of reliable fEPSPs were observed at recording sites located within 450 μm from the stimulation site. Stimulus intensity was related to the number of channels exhibiting fEPSPs. In three mice, we calculated the number of channels that produced detectable fEPSPs responses over a range of stimulus intensities from 5 to 18 μA. We found that the number of responding channels reached a maximum at 16 μA (Figure 1(f)). The effective intensity for inducing 50% of the maximum number of channels was approximately 10 μA. In further experiments, we set the stimulus intensity (typically 9 or 10 μA) to achieve a baseline response of 60%–70% of the maximum.

Figure 1.

The spatial distribution of excitatory synaptic transmission in the RSC was examined using a multielectrode array: (a) Schematic diagram showing the relative positions of the ACC and the RSC. Schematic diagram of an RSC slice placement on the MED64 probe, the blue area is the RSC and the red circle is the stimulated electrode and Schematic diagram of the scale of the electrodes. (b) Light microscopy photograph showed one example for RSC fEPSPs recording by using the MED64 system. Red dotted lines indicate different layers of the RSC; the RSC contains layers I/II/III/V/VI and lacks layer IV; red circle indicates the stimulation site. (c) Sample traces show one channel intensity-dependent fEPSPs (7–11 μA) in RSC. (d, e) All channels fEPSPs were recorded while stimulating channel 21 (indicated by the yellow lightning) with current intensities of 9 and 12 µA. An increase in stimulus intensity resulted in a greater number of channels demonstrating a response, as well as an enhancement in the amplitude of the fEPSPs. The spreading of the fEPSPs displays the network in the RSC. (f) The number of activated channels induced by different stimulation intensities (input–output) in RSC (n = 3 slices/3 mice).

Glutamate receptor-mediated synaptic transmission in the RSC

Glutamate is one of the most widely distributed amino acid transmitters in the brain and is present in most major neural pathways.¹³ Through its regulation of neuronal excitability and metabolic activity in the brain, glutamate acts as an essential transmitter in the regulation of a wide range of physiological and pathological cellular activities. Previous electrophysiological studies have demonstrated that postsynaptic transmission is mediated by glutamatergic AMPA and KA receptors in layer II/III of the ACC in adult rats and mice.^17,25 Due to the close link between the ACC and the RSC, the study of the synaptic transmission in the RSC is important for the mechanisms by which this circuit modulates pain. Therefore, we used the 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 20 μM, 30 min), an AMPA/KA receptor antagonist that is effective in blocking fast synaptic transmission, for bath to determine whether these receptors are the primary mediators of excitatory synaptic transmission in RSC. After bathing with CNQX, almost all fEPSPs in the RSC were blocked (Figure 2(a)–(d)), but not entirely. In the pooled results of seven brain slices, it was found that the response did not completely disappear after bathing CNQX (20 μM), but was reduced to 12.395% ± 2.977% of the baseline (Figure 2(d)), suggesting that all fEPSPs recorded in the RSC are mainly mediated by the action of glutamate on AMPA/KA receptors, with the remaining response perhaps contributed by NMDA receptors. To detect three glutamate receptor-mediated contributions to excitatory synaptic transmission, the NMDA receptor antagonist D-2-amino-5-phosphonovaleric acid (AP-5, 50 μM), the potent AMPA receptor antagonist GYKI 53655 (100 μM) and the AMPA/KA receptor antagonist CNQX (20 μM) were sequentially bath-applied (Figure 3(a)–(d)). After the application of AP-5, fEPSPs were partially inhibited, suggesting a small contribution of NMDA receptors to basal synaptic transmission. And after the application of GYKI 53655, fEPSPs were rapidly and substantially blocked. After final bathing in CNQX, fEPSPs were slightly reduced, but there was no significant difference (AP5: 87.06% ± 2.445% of baseline; GYKI 53655: 11.27% ± 2.329% of baseline; CNQX: 9.793% ± 2.641% of baseline; n = 6 slices/6 mice; Figure 3(c)). Collectively, these results suggest that glutamatergic excitatory synaptic transmission in the RSC is mediated primarily by AMPA receptors, with a small contribution from NMDA receptor.

Figure 2.

Glutamate-mediated synaptic transmission in the RSC: (a) One sample of the fEPSPs of all channels before and after bath CNQX (20 μM). Channel 37 was stimulated with 10 μA (yellow lightning). (b) The outcome of a single channel in the CNQX experiment is presented. The inset traces illustrate representative fEPSPs at the specific time points denoted by the numerical indicators in the graph show that CNQX blocked the potential. (c) The pooled fEPSPs slope suggests that CNQX blocked the potentials of all 18 activated channels in the same slice. (d) Summary result of in all CNQX experiments (n = 7 slices/7 mice).

Figure 3.

AMPA receptors contribute to synaptic transmission in the RSC: (a) The figure shows the results for a single channel with the perfusion of D-AP5 (50 μM), GYKI 53655 (100 μM), CNQX (20 μM) sequentially of the experiment. The insets depict representative fEPSPs at specific time points indicated by the numerical indicators in the figure. (b) Summary results for 10 representative activation channels in one slice. (c) Summary result of in all D-AP5 (50 μM), GYKI 53655 (100 μM), and CNQX (20 μM) experiments (n = 55 channels/6 slices/6 mice). (d) Statistical results all channels of fEPSPs with the perfusion of D-AP5 (50 μM), GYKI 53655 (100 μM), and CNQX (20 μM) sequentially (n = 6 slices/6 mice, unpaired t-test).

The role of N-type VGCCs in RSC excitatory synaptic transmission

We stimulated one channel in the RSC and observed a wide range of responses (Figure 4(a)). Most channels showed a large reduction after the application of the N-type VGCCs blocker ω-Ctx GVIA (1 μM, 30 min) (Figure 4(c)–(e)). For example, all three channels, Ch11, Ch19, and Ch35, experienced reductions, but not to the same extent (Ch11: 52.9% of baseline, Ch19: 31.7% of baseline, Ch35: 85.7% of baseline, Figure 4(b)). To determine whether the contribution of N-type calcium channels to synaptic transmission is not homogeneous in spatial distribution, we categorized the 86 channels in ten slices into three categories according to the degree of reduction from 0%–25%, 25%–50%, and 50%+ of baseline at recording end (Figure 4(f)). The proportions of reductions relative to baseline for the three categories were 54.65%, 32.56%, and 12.79%, respectively. This suggests that the contribution of N-type VGCCs to synaptic transmission in the RSC is heterogeneous.

Figure 4.

N-type VGCCs in RSC excitatory synaptic transmission: (a) Samples of fEPSPs from all channels before and after bathing in ω-Ctx GVIA (1 μM). Channel 20 was stimulated with 9 μA (yellow flash). (b) Three channels with different levels of inhibition after applying ω-Ctx GVIA (1 μM) to a single slice for 30 min. Three different types of samples fEPSPs records acquired at the times indicated by the corresponding numbers are shown on the right side of the figure. (c) Average data for 10 activated channels in a slice. (d, e) Single and average fEPSPs slopes for 86 channels after ω-Ctx GVIA bathing. (f) Scale diagrams categorizing the degree of reduction of 86 channels after ω-Ctx GVIA TFA baths.

To determine the composition of the remaining reactions inhibited by ω-Ctx GVIA, we have applied Cd²⁺ (a potent blocker of all types of VGCCs that completely blocks synaptic transmission) in RSC. After we bathed with 500 μM CdCl₂ in RSC, we found that all channels were completely blocked (Figure 5(a)). Meanwhile, the remaining synaptic responses were also completely blocked by cadmium after bathing ω-Ctx GVIA 1 μM (Figure 5(b)). This suggests that ω-Ctx GVIA likely acts on presynaptic calcium channels to attenuate synaptic strength (Figure 6).

Figure 5.

Roles of other VGCCs in RSC synaptic transmission: (a) The figure shows the results after bathing in cadmium (n = 5 slices/5 mice). The insets depict representative fEPSPs at specific time points indicated by the numerical indicators in the figure. (b) The graph shows the results after sequential bathing of ω-Ctx GVIA and Cd²⁺ (n = 5 slices/5 mice). The insets depict representative fEPSPs at specific time points indicated by the values in the graph. (c) Pooled data of 5 mice (105.1% ± 2.5% of baseline, n = 41 chs/5 slices/5 mice), showing the effect of applying ω-Aga IVA (1 μM) for 20 min. (d) Pooled data of 6 mice (103.4% ± 1.3% of baseline, n = 69 chs/6 slices/6 mice), showing the effect of applying SNX-482 (0.5 μM) for 20 min. (e) Pooled data of 11 mice (100.0% ± 1.7% of baseline, n = 106 chs/11 slices/11 mice), showing the effect of applying Ni²⁺ (100 μM) for 20 min. (f) Pooled data of 16 mice (103.5% ± 0.9% of baseline, n = 106 chs/12 slices/12 mice), showing the effect of applying nimodipine (100 μM) for 30 min. All drugs infused had no effect on synaptic transmission in RSC. The horizontal bars indicate the period of drug application.

Figure 6.

A simplified model to explain N-type VGCCs-mediated excitatory synaptic transmission in RSC. Excitatory transmission in the RSC is mediated by glutamate receptors. Presynaptic N-VGCCs mainly contribute to neurotransmitter release in the RSC. Activation of presynaptic N-VGCCs induces calcium influx. Then, the presynaptic release of excitatory neurotransmitter glutamate is enhanced. NMDA receptors and AMPA receptors activated by glutamate leads to postsynaptic excitatory responses. Postsynaptic AMPA receptors play a dominating role in the synaptic transmission.

The roles of other VGCCs in RSC synaptic transmission

Next, we tested the effects of other VGCCs blockers on RSC glutamatergic synaptic transmission. The P/Q-type VGCCs blocker ω-Aga IVA (1 μM, 20 min) had no effect at all. The response level was 105.1% ± 2.5% at the end of drug treatment and 96.7% ± 2.3% at the end of recording (n = 41 chs/5 slices/5 mice; Figure 5(c)). This is different from previous reports that P/Q-type VGCCs play a dominant role in excitatory synaptic transmission in the hippocampus^26–28 and cerebellum,²⁹ but is consistent with what has been reported for the ACC.¹⁹ It has been shown that R-type VGCCs are involved in excitatory synaptic transmission in rat hippocampus.³⁰ We applied SNX-482 (0.5 μM for 20 min) by bath application to investigate the involvement of R-type VGCCs in RSC. We found that SNX-482 did not produce any significant effect. The response level was 96.0% ± 1.1% at the end of drug treatment and 103.4% ± 1.3% at the end of recording, (n = 69 chs/6 slices/6 mice; Figure 5(d)). Subsequently, we investigated the potential role of low-voltage activation of T-type VGCC by administering the T-type VGCCs inhibitor NiCl₂ (100 μM for 20 min). The response level was 94.4% ± 1.6% at the end of drug treatment and 100.0% ± 1.7% at the end of recording, (n = 106 chs/11 slices/11 mice; Figure 5(e)). Finally, we tested if the L-type VGCCs mediate the RSC synaptic transmission. The response level was 101.8% ± 1.8% at the end of drug treatment and 103.5% ± 0.9% at the end of recording, (n = 106 chs/12 slices/12 mice; Figure 5(f)). The results showed that, with the exception of ω-Ctx GVIA, none of the other four types of calcium channel blockers had a significant effect. In summary, N-type VGCCs are the primary channels implicated in excitatory synaptic transmission with RSC. The other VGCCs that were examined did not demonstrate significant involvement in this process.

Discussion

In this study, we demonstrate for the first time using the MED64 system that excitatory synaptic transmission in the RSC is mediated by glutamate receptors, with AMPA receptors contributing the majority and NMDA receptors contributing a small portion. We found that N-type VGCCs play a dominant role in RSC synaptic transmission, whereas other VGCCs have few effects on synaptic transmission. Moreover, the actions effect of N-type VGCCs in RSC was inhomogeneous in spatial distribution. Future studies using single cell patch-clamp recording will be carried out to characterize other potential VGCCs in excitatory synaptic transmission in the RSC.

Functions of RSC brain regions

Previous studies have consistently shown that the RSC is involved in sensorimotor and cognitive functions.^{1 –5} It contains neurons such as head direction cells, border cells, as well as other cells supporting spatial and contextual encoding.³¹ Functional studies have indicated that the role of the RSC in spatial cognitive processes is multifaceted and includes spatial navigation, orientation, and spatial memory.^5,32 RSC can also be involved in nociceptive modulation. It has been shown that noxious stimulation activates RSC neurons.^6,33,34 The ACC is located in front of the RSC (Figure 1(a)), and plays an important role in the modulation of injury perception and pain-related negative emotions.^9,10 Recent studies have found that migraine and neuropathic pain can lead to enhanced functional connectivity of the RSC with the ACC.^7,8,35 In addition, our recent studies demonstrate that RSC neurons are sending direct projections to ACC neurons; and that this innervation is excitatory and mediated by postsynaptic AMPA receptors.¹¹ Future studies are clearly needed to investigate the potential roles of RSC and RSC-ACC in different types of chronic pain and its related emotional disorders.

Glutamate-mediated synaptic transmission

Glutamate receptors mediate most of the rapid excitatory synaptic transmission in the vertebrate central nervous system, acting on NMDA receptors and non-NMDA receptors.^36,37 Previous electrophysiological studies have shown that in adult rat and mouse ACC, excitatory postsynaptic transmission is mainly mediated by glutamatergic AMPA receptors and KA receptors in layer II/III.^17,25 The insular cortex (IC) also plays an important role in pain processing,³⁸ KA subtype receptors contribute to fast excitatory synaptic transmission in the IC neurons of adult rodents.³⁹ In the hippocampus, synaptically released glutamate in mossy fibers activates postsynaptic KA receptors in addition to AMPA receptors and NMDA receptors.⁴⁰ Importantly, synapses projected from ACC to ACC were also found mixed AMPA receptors and KA receptors.⁴¹ In the present study, we found that excitatory synaptic transmission in RSC is also dominated by glutamate, with AMPA receptors accounting for the major contribution and NMDA receptors make a limited contribution. Although the contribution of KA receptors was not statistically found, the mean value of fEPSPs slope decreased from 11.27% at baseline to 9.79% at baseline after the addition of CNQX. We cannot rule out the possible contribution of some postsynaptic KA receptors in synaptic transmission, since the field recording of EPSPs are not sensitive to detect small currents.

Different types of VGCCs in RSC

N- and P/Q-type calcium channels are the major VGCCs for Ca²⁺ influx to initiate the fast release of neurotransmitters/neuromodulators such as glutamate, acetylcholine, and GABA.^42-45 In ACC, the N-type VGCCs blocker ω-Ctx GVIA significantly reduced glutamatergic synaptic transmission, other VGCCs blockers did not significantly inhibit synaptic responses.¹⁹ This previous finding is in consistent with our current findings in the RSC of adult mice. However, there are reports of other types of VGCCs play roles in synaptic transmission in different regions (Table 1). In the hippocampus, synaptic transmission is regulated by both N- and P/Q-type VGCCs.^26,28,46 In the cerebellum, ω-Ctx GVIA and ω-Aga IVA together virtually eliminated synaptic transmission.²⁹ In mammalian peripheral sympathetic nerves, N-type VGCCs are essential for the transmission of Ca²⁺ influx. P-type and Q-type VGCCs promote Ca²⁺ entry in the presence of N-type VGCCs blocker and high-frequency nerve stimulation.⁴⁷ Our results show that N-type VGCCs blockers reduced glutamatergic synaptic transmission in RSC. Since excitatory synaptic responses in the RSC are mainly mediated by AMPA receptors, the inhibitory effect induced by ω-Ctx GVIA is due to a reduction in presynaptic glutamate release (Figure 6). The current findings show other VGCC blockers did not significantly inhibit RSC synaptic response. Since the drug concentrations used in this experiment were not low compared to other studies, it is highly unlikely that a low dose of the drug would lead to negative results.^19,28,48,49 A possible explanation for this is that other VGCCs may perform some other function than mediating excitatory synaptic transmission. For example, the L-type VGCCs is involved in low frequency stimulation-induced long-term depression in the ACC.^20,25 Taken together, these findings suggest that synapses in the different regions may rely on different types of VGCCs to mediate excitatory synaptic transmission, and that different types of VGCCs mediate excitatory synaptic transmission to different extents. The ω-Ctx GVIA blocked about 50% of total synaptic transmission in the ACC,¹⁹ but blocked about 25% of total synaptic transmission in the RSC. The remaining response can be completely blocked by Cd²⁺. Therefore, future studies are still needed to identify the receptors or channels that mediate the remaining 75% of RSC synaptic responses under basal conditions.

Table 1.

The role of VGCCs mediate synaptic transmission in different areas.

Area	Types of VGCCs	References
RSC	N	Present study
ACC	N	Kang et al.¹⁹
Hippocampus	N	Wheeler et al.²⁸
	P/Q	Castillo et al.³⁴
	P/Q	Luebke et al.²⁶
	R	Gasparini et al.³⁰
Cerebellum	NP/Q	Regehr and Mintz²⁹
Peripheral regions	NP/Q	Wright and Angus³⁵
Spinal cord	NP/Q	Takahashi and Momiyama²⁷

In summary, our present study demonstrates for the first time that excitatory synaptic transmission in adult mouse RSC is mediated by glutamate, and that AMPA receptors make a major contribution. We also demonstrated the importance of N-type VGCCs in mediating excitatory synaptic transmission in adult mice RSC. Future studies using single cell patch-clamp recording and behavioral tests will be carried out to characterize the roles of other glutamate receptors and potential VGCCs in excitatory synaptic transmission in the RSC.

Footnotes

This work was supported by funding to M.Z. from Oujiang Laboratory (OJQD2022004).

Author contributions

J.J.W.,Y.J.M. and M.Z. designed the research plan. J.J.W. and Y.J.M. performed the research. J.J.W. and M.Z. wrote the manuscript. All authors discussed the manuscript. All authors read and approved the final manuscript.

Declaration of conflicting interests

The author(s) declared no potential conflicts of interest with respect to the research,authorship,and/or publication of this article.

Funding

The author(s) received no financial support for the research,authorship,and/or publication of this article.

Ethics statement

Experiments research protocols have been approved by Experimental Animal Ethics Review Committee at the Oujiang Laboratory.

ORCID iD

Min Zhuo

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Characterization of excitatory synaptic transmission in the retrosplenial cortex of adult mice

Abstract

Keywords

Introduction

Methods

Animals

Brain slice preparation

Preparation of the multielectrode array

Field potential recording in adult RSC slices

Data analysis

Drug

Results

Spatial distribution of extracellular fEPSPs in the RSC

Glutamate receptor-mediated synaptic transmission in the RSC

The role of N-type VGCCs in RSC excitatory synaptic transmission

The roles of other VGCCs in RSC synaptic transmission

Discussion

Functions of RSC brain regions

Glutamate-mediated synaptic transmission

Different types of VGCCs in RSC

Footnotes

Author contributions

Declaration of conflicting interests

Funding

Ethics statement

ORCID iD

References