Ventral Midbrain NTS1 Receptors Mediate Conditioned Reward Induced by the Neurotensin Analog, D-Tyr[11]neurotensin

Abstract

The present study was aimed at characterizing the mechanisms by which neurotensin (NT) is acting within the ventral midbrain to induce a psychostimulant-like effect. In a first experiment, we determine which subtype(s) of NT receptors is/are involved in the reward-inducing effect of ventral midbrain microinjection of NT using the conditioned place-preference (CPP) paradigm. In a second study, we used in vitro patch clamp recording technique to characterize the NT receptor subtype(s) involved in the modulation of glutamatergic neurotransmission (excitatory post-synaptic current, EPSC) in ventral tegmental neurons that expressed ([Formula: see text]), or do not express ([Formula: see text]), a hyperpolarization-activated cationic current. Behavioral studies were performed with adult male Long-Evans rats while electrophysiological recordings were obtained from brain slices of rat pups aged between 14 and 21 days. Results show that bilateral ventral midbrain microinjections of 1.5 and 3 nmol of D-Tyr[(11)]NT induced a CPP that was respectively attenuated or blocked by co-injection with 1.2 nmol of the NTS1/NTS2 antagonist, SR142948, and the preferred NTS1 antagonist, SR48692. In electrophysiological experiments, D-Tyr[(11)]NT (0.01-0.5 μM) attenuated glutamatergic EPSC in [Formula: see text] but enhanced it in [Formula: see text] neurons. The attenuation effect ([Formula: see text] neurons) was blocked by SR142948 (0.1 μM) while the enhancement effect ([Formula: see text] neurons) was blocked by both antagonists (0.1 μM). These findings suggest that (i) NT is acting on ventral midbrain NTS1 receptors to induce a rewarding effect and (ii) that this psychostimulant-like effect could be due to a direct action of NT on dopamine neurons and/or an enhancement of glutamatergic inputs to non-dopamine ([Formula: see text]) neurons.

Keywords: conditioned reward; glutamate; neurotensin; ventral midbrain.

Figure 1

Induction of a CPP by [D-Tyr¹¹]NT. Top panel illustrates the preference score measured on the test day for the animals that were injected with 1.5 nmol (NT1.5, n = 6), 3 nmol (NT3, n = 13) of [D-Tyr¹¹]NT or its vehicle (VEH, n = 15). Preference score corresponds to the amount of time (in sec) spent in the paired compartment on the test day minus the time spent at baseline in the same compartment. Measures of locomotor activity recorded during the preference test for the animals in each treatment group are presented in the middle panel (horizontal) and bottom panel (stereotypy-like). Asterisks indicate a statistical significant difference with VEH (^**p < 0.01; ^***p < 0.001).

Figure 2

Mean preference score measured on the test day in the unpaired (top panel) and neutral (bottom panel) compartment for the animals that were injected with 1.5 nmol (NT1.5, n = 6), 3 nmol (NT3, n = 13) of [D-Tyr¹¹]NT or its vehicle (VEH, n = 15). Preference score corresponds to the amount of time (in sec) spent in the unpaired or neutral compartment on the test day minus the time spent at baseline in the same compartment. See text for details.

Figure 3

Effects of SR142948 and SR48692 on [D-Tyr¹¹]NT-induced CPP. Top panel illustrates the preference score measured on the test day for the animals that were injected with 3 nmol of [D-Tyr¹¹]NT (NT, n = 13), 1.2 nmol of SR142948 (SR142, n = 10), 1.2 nmol of SR48692 (SR48, n = 7), SR142948 + [D-Tyr¹¹]NT (SR142+NT, n = 12), SR48292 + [D-Tyr¹¹]NT (SR48+NT, n = 10) or the vehicle (VEH, n = 15). Preference score corresponds to the amount of time (in sec) spent in the paired compartment on the test day minus the time spent at baseline in the same compartment. Measures of locomotor activity recorded during the preference test for the animals in each treatment group are presented in the middle panel (horizontal) and bottom panel (stereotypy-like). The asterisks and the cross indicate a statistical significant difference with VEH (^*p < 0.01; ^***p < 0.001) and NT (⁺p < 0.05) respectively.

Figure 4

Effect of [D-Tyr¹¹]NT and antagonists on ${\text{I}}_{\text{h}}^{+}$ neurons. Panel 1: Current traces of glutamatergic EPSC recorded during superfusion of [D-Tyr¹¹]NT; control (1), with [D-Tyr¹¹]NT (0.01 μM) (2) and following the washout of [D-Tyr¹¹]NT (3) at a holding membrane potential of −70 mV in ${\text{I}}_{\text{h}}^{+}$ neurons (n = 6). Panel 2: Current traces of glutamatergic EPSC recorded during superfusion with SR142948 (0.1 μM) (1), with SR142948 and [D-Tyr¹¹]NT (0.01 μM) (2) and with SR142948 following the washout of (3) at a holding membrane potential of −70 mV in ${\text{I}}_{\text{h}}^{+}$ neurons (n = 5). Panel 3: Current traces of glutamatergic EPSC recorded during superfusion with SR48692 (0.1 μM) (1), with SR48692 and [D-Tyr¹¹]NT (0.01 μM) (2) and with SR48692 following the washout of [D-Tyr¹¹]NT (3) at a holding membrane potential of −70 mV in ${\text{I}}_{\text{h}}^{+}$ neurons (n = 4).

Figure 5

Effects of [D-Tyr¹¹]NT on glutamatergic EPSCs in ${\text{I}}_{\text{h}}^{+}$ and ${\text{I}}_{\text{h}}^{-}$ neurons. Mean percent change in EPSC amplitude recorded in ${\text{I}}_{\text{h}}^{+}$ (black bar) and ${\text{I}}_{\text{h}}^{-}$ (white bar) following application of different concentrations of [D-Tyr¹¹]NT. The number of neurons recorded at each concentration is as follow: 0.01 μM, n = 12 (${\text{I}}_{\text{h}}^{+}$ n = 6, ${\text{I}}_{\text{h}}^{-}$ n = 6); 0.1 μM, n = 12 (${\text{I}}_{\text{h}}^{+}$ n = 6, ${\text{I}}_{\text{h}}^{-}$ n = 6); 0.5 μM, n = 16 (${\text{I}}_{\text{h}}^{+}$ n = 7, ${\text{I}}_{\text{h}}^{-}$ n = 9). All concentrations of D-Tyr[11]NT are reported in μM. Asterisks indicate a statistically significant difference with the lowest concentration (^**p < 0.05; ^***p < 0.001). See text for details.

Figure 6

Effect of [D-Tyr¹¹]NT and antagonists on ${\text{I}}_{\text{h}}^{-}$ neurons. Panel 1: Current traces of glutamatergic EPSC recorded during superfusion of [D-Tyr¹¹]NT; control (1), with [D-Tyr¹¹]NT (0.01 μM) (2) and following the washout of [D-Tyr¹¹]NT (3) at a holding membrane potential of −70 mV in ${\text{I}}_{\text{h}}^{-}$ neurons (n = 6). Panel 2: Current traces of glutamatergic EPSC recorded during superfusion with SR142948 (0.1 μM) (1), with SR142948 and [D-Tyr¹¹]NT (0.01 μM) (2) and with SR142948 following the washout of [D-Tyr¹¹]NT (3) at a holding membrane potential of −70 mV in ${\text{I}}_{\text{h}}^{-}$ neurons (n = 5). Panel 3: Current traces of glutamatergic EPSC recorded during superfusion with SR48692 (0.1 μM) (1), with SR48692 and [D-Tyr¹¹]NT (0.01 μM) (2) and with SR48692 following the washout of [D-Tyr¹¹]NT (3) at a holding membrane potential of −70 mV in ${\text{I}}_{\text{h}}^{-}$ neurons (n = 5).

Figure 7

Effect of SR142948 and SR48692 on glutamatergic EPSCs in ${\text{I}}_{\text{h}}^{+}$ and ${\text{I}}_{\text{h}}^{-}$ neurons. Mean percent change in EPSC amplitude recorded in ${\text{I}}_{\text{h}}^{+}$ (A) and ${\text{I}}_{\text{h}}^{-}$ neurons (B) following application of [D-Tyr¹¹]NT alone or in the presence of SR142948 or SR48692. The number of neurons recorded under each condition is as follow: ${\text{I}}_{\text{h}}^{+}$ neurons, [D-Tyr¹¹]NT (0.01 μM, n = 6; 0.1 μM, n = 6); SR142948 (0.1 μM, n = 5; 0.5 μM, n = 4); SR48692 (0.1 μM, n = 4; 0.5 μM, n = 4); ${\text{I}}_{\text{h}}^{-}$ neurons, [D-Tyr¹¹]NT (0.01 μM, n = 6; 0.1 μM, n = 6); SR142948 (0.1 μM, n = 5; 0.5 μM, n = 5); SR48692 (0.1 μM, n = 5; 0.5 μM, n = 4). All concentrations of D-Tyr[11]NT, SR142948, and SR48692 are reported in μM. Asterisks and crosses indicate a statistically significant difference with [D-Tyr¹¹]NT alone at 0.01 μM and 0.1 μM respectively (^***p < 0.001 with 0.01 μM; ⁺⁺⁺p < 0.001 with 0.1 μM). The ^### sign indicates a statistical significant difference between SR142948 (0.5 μM) +D-Tyr[11]NT (0.1 μM) and SR48692 (0.5 μM) +D-Tyr[11]NT (0.1 μM). See text for details.