Inhibition by opioids acting on μ‐receptors of GABAergic and glutamatergic postsynaptic potentials in single rat periaqueductal gray neurones in vitro

1 Membrane properties of rat periaqueductal gray neurones were investigated by use of intracellular recordings from single neurones in brain slices. Morphological properties and anatomical location of each impaled neurone were characterized by intracellular staining with biocytin. The present paper considers the properties of electrically‐evoked and spontaneous postsynaptic potentials impinging on periaqueductal gray neurones, and the actions of opioids on postsynaptic potentials in neurones which were not directly hyperpolarized by opioids. The preceding paper considers neurones which were hyperpolarized by opioids. 2 Electrical stimulation in the vicinity of impaled neurones evoked postsynaptic potentials having fast (duration at half‐maximal amplitude 37 ± 2 ms, n = 65) and in some cases slow (duration at half‐maximal amplitude 817 ± 187 ms, n = 3) components. Amplitudes of evoked potentials were dependent on stimulus voltage, membrane potential, and were abolished during superfusion with solutions containing tetrodoxotoxin (100 nm to 1 μm, n = 5) or Co2+ (4 mm, n = 2). 3 Fast postsynaptic potentials were mediated predominantly by activation of glutamate and GABAA receptors. The GABAA‐receptor antagonist, bicucuilline (30 μm), inhibited postsynaptic potentials by 44 ± 8% (n = 14). The non‐NMDA‐receptor antagonist, 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione (10 μm), inhibited postsynaptic potentials by 48 ± 6% (n = 16). Combined superfusion of bicuculline (30 μm) and 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione (10 μm) inhibited postsynaptic potentials by 93 ± 1% (n = 8). Additional superfusion of the NMDA‐receptor antagonist, (±)‐2‐amino‐5‐phosphonovaleric acid (50 μm) inhibited synaptic potentials by 94 ± 1% (n = 3). 4 Slow inhibitory postsynaptic potentials were observed in 12% (8/65) of neurones. They reversed polarity between − 100 and − 110 mV, and were abolished by superfusion with spiperone (1 μm, n = 2), but not the α2‐antagonist, idazoxan (3 μm, n = 2). 5 Selective μ‐receptor agonists inhibited fast postsynaptic potentials in all neurones tested which were not directly hyperpolarized by opioids. Met‐enkephalin (30 μm) and Tyr‐d‐Ala‐Gly‐MePhe‐Glyol (3 μm) inhibited postsynaptic potentials by 53 ± 3% and 49 ± 3%, respectively. This effect was completely antagonised by naloxone (1 μm, n = 3). A small inhibition produced by the selective δ‐receptor agonist, Tyr‐d‐Pen‐Gly‐Phe‐d‐Pen‐enkephalin (3 μm, 26 ± 4%, n = 14), was antagonized by naloxone (1 μm), but not by the selective δ‐receptor antagonist, naltrindole (10 nm), suggesting non‐specific μ‐receptor activation by this agonist. The selective κ‐receptor agonist, U50488H (3 μm), also consistently inhibited postsynaptic potentials by 45 ± 15% (n = 4). However, this effect was not fully reversed by naloxone (1 μm) suggesting a non‐specific action. 6 Both glutamatergic and GABAergic components of fast postsynaptic potentials were inhibited by Met‐enkephalin (10 or 30 μm). Met‐enkephalin inhibited postsynaptic potentials by 55 ± 5% (n = 12) in the presence of 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione (10 μm, predominantly GABAergic component). Met‐enkephalin did not affect the response to GABA applied directly by pressure ejection, indicating that opioids exclusively inhibited presynaptic release of GABA. Met‐enkephalin (10–30 μm) inhibited postsynaptic potentials by 48 ± 6% (n = 11) in the presence of bicuculline (30 μm, predominantly glutamatergic component). In the presence of both bicuculline and 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione, Met‐enkephalin inhibited the small residual component of the synaptic potential by 42 ± 15% (n = 2). 7 Frequent spontaneous synaptic potentials were also observed in 11% (10/94) of the neurones which were not directly hyperpolarized by opioids. These were reversibly abolished by bicuculline (30 μm, n = 5) and substantially inhibited by Met‐enkephalin (30 μm, n = 6), but were unaffected by 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione (10 μm, n = 2). 8 In conclusion, fast glutamatergic and GABAergic synaptic potentials were evoked by electrical stimulation throughout the lateral and ventrolateral periaqueductal gray. Slow inhibitory synaptic potentials were also evoked in some neurones. Opioids acting on μ‐receptors inhibited both GABAergic and glutamatergic components of synaptic potentials throughout this brain region.