Presynaptic GABAB receptors reduce transmission at parabrachial synapses in the lateral central amygdala by inhibiting N-type calcium channels

Abstract

The nocioceptive information carried by neurons of the pontine parabrachial nucleus to neurons of the lateral division of the central amydala (CeA-L) is thought to contribute to the affective components of pain and is required for the formation of conditioned-fear memories. Importantly, excitatory transmission between parabrachial axon terminals and CeA-L neurons can be inhibited by a number of presynaptic receptors linked to Gi/o-type G-proteins, including α2-adrenoceptors and GABAB receptors. While the intracellular signalling pathway responsible for α2-adrenoceptor inhibition of synaptic transmission at this synapse is known, the mechanism by which GABAB receptors inhibits transmission has not been determined. The present study demonstrates that activation of presynaptic GABAB receptors reduces excitatory transmission between parabrachial axon terminals and CeA-L neurons by inhibiting N-type calcium channels. While the involvement of Gβγ subunits in mediating the inhibitory effects of GABAB receptors on N-type calcium channels is unclear, this inhibition does not involve Gβγ-independent activation of pp60C-src tyrosine kinase. The results of this study further enhance our understanding of the modulation of the excitatory input from parabrachial axon terminals to CeA-L neurons and indicate that presynaptic GABAB receptors at this synapse could be valuable therapeutic targets for the treatment of fear- and pain-related disorders.

Figure 1. GABA_B-receptor and noradrenenergic inhibition of parabrachial-CeA-L EPSCs is blocked by pre-incubation with 50 μM N-ethylmaleimide (NEM) for 10 minutes prior to application of baclofen or noradrenaline.

(A) Parabrachial-CeA-L EPSC amplitudes during the application of 2 μM baclofen (bacl); inset - average of control and baclofen responses. (B) Parabrachial-CeA-L EPSC amplitudes during the application of 10 μM noradrenaline (NAd); inset - average of control and noradrenaline responses. (C,D) Parabrachial-CeA-L EPSC amplitudes in control, and with subsequent application of NEM followed by baclofen (bacl) (C) and noradrenaline (NAd) (D). Inset – average traces in NEM (NEM) and NEM + baclofen (NEM+ bacl) or noradrenaline (NEM + NAd) shown.

Figure 2. GABA_B-mediated inhibition does not require G_βγ signalling.

(A) Parabrachial-CeA-L EPSC amplitude recorded from synapses loaded with the myristlated compound mSIRK in control conditions and after the addition of noradrenaline (NAd) and baclofen (bacl). Below - average EPSCs in each condition. (B) Baclofen (bacl) activated post-synaptic currents recorded from CeA-L cells in control conditions and after mSIRK application.

Figure 3. GABA_B-mediated inhibition of parabrachial-CeA-L EPSCs by baclofen is reduced by blocking N-type calcium channels but not by pre-incubation with the selective src-kinase inhibitor PP2.

(A) Parabrachial-CeA-L EPSC amplitudes recorded from synapses in control conditions and following sequential addition of ω-conotoxin CVID (CIVD; 200 nM) and baclofen (bacl; 2 μM). (B) Parabrachial-CeA-L EPSC amplitudes recorded from synapses in slices pre-incubated for 90 min in PP2 in the absence and presence of baclofen. Average EPSCs in control and baclofen are shown on right.