Presynaptic opioid receptors on noradrenergic and serotonergic neurons in the human as compared to the rat neocortex

Abstract

1. Electrically evoked release of [3H]-noradrenaline ([3H]-NA) or [3H]-5-hydroxytryptamine ([3H]-5-HT) in slices of human and the rat neocortex was used to characterize presynaptic opioid receptors. 2. Release of [3H]-NA in rat neocortical slices was reduced only by the mu-receptor agonist DAMGO (pIC50: 7.27, CI95: [7.22, 7.32]; Imax: 77.6+/-1.6%; antagonized by naloxone: pA2: 8.88, CI95: [8.78, 8.98]). 3. Release of [3H]-NA in human neocortical slices was unaffected by DAMGO, but inhibited by the delta-receptor agonist DPDPE (Imax: 25.7+/-2.2%) and the kappa-receptor agonist U-50,488H (19.7+/-2.7% inhibition at 1 microM). Both effects were antagonized by naltrindole (1 microM). 4. Release of [3H]-5-HT in rat neocortical slices, was inhibited by DAMGO (10 microM) and U-50,488H (1 and 10 microM) only in the presence of the 5-HT receptor antagonist methiotepin (1 microM). 5. Release of [3H]-5-HT in human neocortical slices was unaffected by DPDPE, but U-50,488H (Imax: 40.8+/-8.3%; antagonized by 0.1 microM norbinaltorphimine) and DAMGO (16.4+/-3.9% inhibition at 1 microM; antagonized by 0.1 microM naloxone) acted inhibitory. 6. Release of [3H]-5-HT in human neocortical slices was reduced by nociceptin/orphanin (0.1 and 1 microM). These effects were antagonized by the ORL1 antagonist J-113397 (1-[(3R,4R)-1-cyclo-octylmethyl-3-hydroxymethyl-4-piperidyl]-3-ethyl-1,3-dihydro-2H-benzimidazol-2-one; 0.1 microM). 7. This study provides evidence for significant species differences in opioid receptor-mediated modulation of NA and 5-HT-release in human vs rat neocortex. In rats, mu-opioid receptors modulate NA release, but 5-HT release is only weakly affected by mu- and kappa-opioids. In contrast, NA release in human neocortex is modulated via delta-opioid receptors, but 5-HT release mainly via kappa-opioid receptors. In addition also the ORL1 receptor seems to be involved in 5-HT release modulation.

Figure 1

Time course of ³H-outflow from human neocortical slices preincubated with either 0.1 μM [³H]-NA (a), or 0.1 μM [³H]-5-HT (b): effects of Ca²⁺-free medium and tetrodotoxin (TTX, 0.3 μM). During superfusion ((a): in the presence of desipramine, 1 μM; (b): in the presence of 6-NQ and nomifensin, 1 μM each), overflow of [³H] was evoked by two electrical field stimulations ((a): 8 pulses at 100 Hz, 2 ms, 34 mA and (b): 2 × 8 pulses at 100 Hz, 2 ms, 65 mA) as indicated by the black squares (S₁, S₂) in the lowest plots. Ca²⁺-free or TTX-containing media were added 8 min before S₂, as shown by the horizontal bars; n=4–5 for each curve.

Figure 2

Effects of DAMGO on electrically-evoked overflow of [³H] in rat neocortical slices preincubated with 0.1 μM [³H]-NA. Following preincubation the slices were superfused either in presence of desipramine (1 μM) alone, or in the additional presence of naloxone (0.1 μM). During superfusion, the overflow of [³H] was evoked by three electrical field stimulations (S₁, S₂, S₃; 4 pulses at 100 Hz, 2 ms, 34 mA). DAMGO was added in increasing concentrations 8 min before S₂ or S₃, respectively, as shown on the abscissa. Effects of DAMGO on the evoked overflow of [³H] are shown as S_n/S₁ ratios expressed as percentage of the corresponding untreated controls; means±s.e.m.; n=8–18, per drug concentration.

Figure 3

Effects of DAMGO, U-50,488H and DPDPE on electrically evoked overflow of [³H] in human neocortical slices preincubated with 0.1 μM [³H]-NA. Following preincubation the slices were superfused either in presence of desipramine (1 μM) alone, or in the additional presence of naltrindole (1 μM). During superfusion, overflow of [³H] was evoked by three electrical field stimulations (S₁, S₂, S₃; 8 pulses at 100 Hz, 2 ms, 34 mA). The opioid receptor agonists were added in increasing concentrations 8 min before S₂ or S₃, respectively, as shown on the abscissa. Effects of drugs on the evoked overflow of [³H] are shown as S_n/S₁ ratios expressed as percentage of the corresponding untreated controls (dotted line). Significance of differences vs corresponding controls: ^**P<0.01, ^***P<0.001; vs absence of naltrindole: ⁺P<0.05, ⁺⁺⁺P<0.001; means±s.e.m.; n=7–10, per drug concentration (data from 3 to 4 patients; for power analysis: see text).

Figure 4

Effects of DAMGO, U-50,488H and DPDPE on electrically evoked overflow of [³H] in rat neocortical slices preincubated with 0.1 μM [³H]-5-HT (in the presence of 1 μM nomifensin). Following preincubation, the slices were superfused either in the presence of 6-nitroquipazine and nomifensin (1 μM, each), or in the additional presence of methiotepin (1 μM). During superfusion, overflow of [³H] was evoked by three electrical field stimulations (S₁, S₂, S₃; 2 × 4 pulses at 100 Hz, 2 ms, 65 mA). The opioid receptor agonists were added in increasing concentrations 8 min before S₂ or S₃, respectively, as shown on the abscissa. Effects of drugs on the evoked overflow of [³H] are shown as S_n/S₁ ratios expressed as percentage of the corresponding untreated controls (dotted line); means±s.e.m.; n=5–15 per drug concentration and condition. In the absence of methiotepin throughout, none of the drug effects differed significantly from the corresponding controls; significance of drug effects (in presence of methiotepin) vs corresponding controls: ^*P<0.05; ^**P<0.01.

Figure 5

Effects of DAMGO and DPDPE on electrically evoked overflow of [³H] in human neocortical slices preincubated with 0.1 μM [³H]-5-HT (in the presence of 1 μM nomifensin). Following preincubation, the slices were superfused either in presence of 6-NQ and nomifensin (1 μM, each, hatched columns), or in the additional presence of nBNI (0.01 μM, grey columns) or of naloxone (0.1 μM, black columns) throughout superfusion. During superfusion, overflow of [³H] was evoked by three electrical field stimulations (S₁, S₂, S₃; 2 × 8 pulses at 100 Hz, 2 ms, 65 mA). The opioid receptor agonists were added in increasing concentrations 8 min before S₂ or S₃, respectively, as shown on the abscissa. Effects of drugs on the evoked overflow of [³H] are shown as S_n/S₁ ratios expressed as percentage of the corresponding untreated controls (dotted line). Significance of effects vs corresponding controls: ^**P<0.01; means±s.e.m.; n=9–22 per drug concentration (data from 2 to 3 patients; for power analysis: see text).

Figure 6

Effects of U-50,488H on electrically evoked overflow of [³H] in human neocortical slices preincubated with 0.1 μM [³H]-5-HT (in the presence of 1 μM nomifensin). Following preincubation, the slices were superfused either in presence of 6-NQ and nomifensin (1 μM, each, filled circles), or in the additional presence of nBNI (0.01 μM, open circles) throughout superfusion. During superfusion, overflow of [³H] was evoked by three electrical field stimulations (S₁, S₂, S₃; 2 × 8 pulses at 100 Hz, 2 ms, 65 mA). U-50,488H was added in increasing concentrations 8 min before S₂ or S₃, respectively, as shown on the abscissa. Effects of U-50,488H (in absence or presence of nBNI) on the evoked overflow of [³H] are shown as S_n/S₁ ratios expressed as percentage of the corresponding untreated controls (dotted line). Significance of effects vs corresponding controls: ^*P<0.05, ^**P<0.01; ^***P<0.001; vs U-50,488H alone: ^#P<0.05, ^###P<0.001; means±s.e.m.; n=7–22 per drug concentration (data from 2 to 6 patients; for power analysis: see text).

Figure 7

Effects of nociceptin (N/OFQ) in the absence or presence of the ORL1 antagonist J-113397 (0.1 μM) on the electrically evoked overflow of [³H] from human neocortical slices preincubated with [³H]-5-HT (in the presence of 1 μM nomifensin). Following preincubation, the slices were superfused either in presence of 6-NQ and nomifensin (1 μM, each, gray columns), or in the additional presence of J-113397 (0.1 μM, black columns) throughout superfusion. During superfusion, overflow of [³H] was evoked by three electrical field stimulations (S₁, S₂, S₃; 2 × 8 pulses at 100 Hz, 2 ms, 65 mA). N/OFQ was added in increasing concentrations 8 min before S₂ or S₃, respectively, as shown on the abscissa. Effects of N/OFQ (in absence or presence of J-113397) on the evoked overflow of [³H] are shown as S_n/S₁ ratios expressed as percentage of the corresponding untreated controls (dotted line). Significance of effects vs corresponding controls: ^***P<0.001; significance of effects vs absence of J-113397: ⁺⁺P<0.01; means±s.e.m.; n=12–22 per drug concentration (data from three patients; for power analysis: see text).