The Lateral Preoptic Area: A Novel Regulator of Reward Seeking and Neuronal Activity in the Ventral Tegmental Area

Abstract

The lateral preoptic area (LPO) is a hypothalamic region whose function has been largely unexplored. Its direct and indirect projections to the ventral tegmental area (VTA) suggest that the LPO could modulate the activity of the VTA and the reward-related behaviors that the VTA underlies. We examined the role of the LPO on reward taking and seeking using operant self-administration of cocaine or sucrose. Rats were trained to self-administer cocaine or sucrose and then subjected to extinction, whereby responding was no longer reinforced. We tested if stimulating the LPO pharmacologically with bicuculline or chemogenetically with Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) modifies self-administration and/or seeking. In another set of experiments, we tested if manipulating the LPO influences cocaine self-administration during and after punishment. To examine the functional connectivity between the LPO and VTA, we used in vivo electrophysiology recordings in anesthetized rats. We tested if stimulating the LPO modifies the activity of GABA and dopamine neurons of the VTA. We found that stimulating the LPO reinstated cocaine and sucrose seeking behavior but had no effect on reward intake. Furthermore, both stimulating and inhibiting the LPO prevented the sustained reduction in cocaine intake seen after punishment. Finally, stimulating the LPO inhibited the activity of VTA GABA neurons while enhancing that of VTA dopamine neurons. These findings indicate that the LPO has the capacity to drive reward seeking, modulate sustained reductions in self-administration following punishment, and regulate the activity of VTA neurons. Taken together, these findings implicate the LPO as a previously overlooked member of the reward circuit.

Keywords: cocaine; dopamine; punishment; relapse; reward; self-administration; sucrose.

FIGURE 1

Pharmacological stimulation of the LPO promotes cocaine seeking, but does not change cocaine self-administration. (A) Timeline of behavioral procedures. SA, self-administration; FR: fixed ratio (number of responses required to obtain one cocaine infusion, depicted with pink line). (B) Location of LPO injections for aCSF (gray) and bicuculline (Bic, purple). (C) Cocaine self-administration behavior. There was slightly more responding in the active hole in rats that would later receive bicuculline compared with those that would later receive aCSF; however, during the last 3 days of self-administration that preceded the self-administration test, groups did not differ. (D) Self-administration test (SA test). Stimulating the LPO with bicuculline did not change active hole or inactive hole responding relative to aCSF or the average of the last 3 days of self-administration (SA Pre). (E) Extinction behavior. Both groups extinguished responding on the previously active hole. There was no difference between groups across extinction nor over the last 3 days of extinction (Ext Pre). (F) Extinction test (Ext Test). Stimulating the LPO with bicuculline reinstated cocaine seeking behavior, observed as increased responding on the previously active hole (HSD, ^∗∗P < 0.01) but not inactive hole (HSD, P = 0.47). Symbols are means ± SEM for each group; lines are individual subjects. See main text for detailed statistics.

FIGURE 2

Chemogenetic stimulation of the LPO promotes cocaine seeking. (A) Timeline of behavioral procedures. SA, self-administration; FR, fixed ratio (number of responses required to obtain one cocaine infusion, depicted by pink lines). (B) Representative image of hM3Dq-mCitrine fluorescence in the LPO. (C) Localization of viral expression for GFP (gray) and hM3Dq (purple). (D) Cocaine self-administration behavior. Both groups acquired cocaine self-administration and there was no difference between groups across self-administration or over the last 3 days of self-administration (SA Pre). (E) Extinction behavior. Both groups extinguished responding on the previously active hole. There was no difference between groups across extinction or over the last 3 days of extinction (Ext Pre). (F) Extinction test (Ext Test). In the hM3Dq group, stimulating the LPO with CNO reinstated cocaine seeking behavior, observed as increased responding on the previously active hole (HSD, ^∗∗∗P < 0.001) but not inactive hole (HSD, P = 0.99). Symbols are means ± SEM for each group; lines are individual subjects. See main text for detailed statistics.

FIGURE 3

Pharmacological stimulation of the LPO promotes sucrose seeking, but does not change sucrose self-administration. (A) Timeline of behavioral procedures. SA, self-administration; FR, fixed ratio (number of responses required to obtain one reward delivery, depicted with pink lines). Pellets: number of pellets obtained per reward delivery, depicted with pink lines. (B) Location of LPO injections for aCSF (gray) and bicuculline (Bic, purple). (C) Sucrose self-administration behavior. Both groups acquired sucrose self-administration and there was no difference between groups across self-administration or over the last 3 days of self-administration (SA Pre). Rats updated responding with changes in FR schedule and number of rewards per delivery. (D) Self-administration test (SA Test). Stimulating the LPO with bicuculline did not change active hole or inactive hole responding relative to aCSF controls. (E) Extinction behavior. Both groups extinguished responding on the previously active hole. There was no difference between groups across extinction or over the last 3 days of extinction (Ext Pre). (F) Extinction test (Ext Test). Stimulating the LPO with bicuculline reinstated sucrose seeking behavior, observed as increased responding on the previously active hole (HSD, ^∗∗P < 0.01) but not the inactive hole (HSD, P = 1.00). Symbols are mean ± SEM for each group; lines are individual subjects. See main text for detailed statistics.

FIGURE 4

Pharmacological manipulation of the LPO disrupts the reduction in self-administration of cocaine after punishment. (A) Timeline of behavioral procedures. SA, self-administration; FR, fixed ratio (number of responses required to obtain one cocaine infusion, depicted with pink line). (B) Location of LPO injections for aCSF (gray), bicuculline (Bic, purple), and baclofen + muscimol (Bac + Mus, green). (C) Cocaine self-administration behavior, data are mean ± SEM of each group. There was no difference between groups across self-administration or over the last 3 days of self-administration (SA). (D) Behavior during punishment. Lines are individual subjects; symbols and error bars represent means ± SEM of each group. During footshock (EFS) punishment, all groups decreased the number of infusions relative to pre punishment (SA) (all HSD comparisons, Ps < 0.001), and this occurred to a similar extent in animals receiving aCSF, bicuculline, or baclofen + muscimol. On the day following punishment (Post), only the aCSF group remained significantly below baseline intake (HSD, aCSF: ^∗∗∗P < 0.001), whereas the other groups returned to pre-baseline intake (HSD, Bic: P = 0.20; Bac + Mus: P = 0.99).

FIGURE 5

Pharmacological stimulation of the LPO enhances the firing rate of VTA dopamine neurons and inhibits that of VTA GABA neurons. (A) Location of LPO injections: aCSF (gray), bicuculline (Bic, red for GABA neurons and blue for dopamine neurons), during recordings of GABA neurons (squares) or dopamine neurons (circles). (B) Locations of dopamine (circles) and GABA (squares) neurons within the VTA. Color indicates corresponding intra-LPO injection: aCSF (gray), bicuculline (Bic, red for GABA neurons and blue for dopamine neurons). (C) Firing in GABA neurons (delta from baseline) before and after the administration of aCSF (gray) or bicuculline (Bic, red). Time is relative to onset of 3-min microinjection; each point represents the mean ± SEM values of each group. Stimulating the LPO with bicuculline decreased firing in GABA neurons relative to aCSF control and baseline (pre-injection) activity (group × time interaction: F₍₈,₉₆₎ = 3.29, P = 0.0023, HSD, ^∗P < 0.05 compared with all pre-injection time-points). (D) Representative firing rate in a GABA neuron. There was substantial decrease in firing rate throughout injection and following. (E) Average waveform and recording traces for the neuron shown in graph (D). Symbols denote the time period from which each trace was (obtained. (F) Firing in dopamine neuron (delta from baseline) before and after the administration of aCSF (gray) or bicuculline (Bic, blue). Time is relative to onset of the 3-min microinjection; each point represents the mean ± SEM values of each group. Stimulating the LPO with bicuculline increased the firing rate of dopamine neurons, relative to aCSF control and baseline (pre-injection) activity (group × time interaction: F₍₈,₁₂₂₎ = 2.87, P = 0.0060, HSD, ^∗P < 0.05 compared with all pre-injection time-points). (G) Representative firing rate in a dopamine neuron. There was an increase in firing rate throughout the injection and following. (H) Average waveform and recording traces for the neuron shown in graph (G). Symbols denote the time period from which each trace was obtained. (I) Burst characteristics of dopamine neurons before and after the administration of aCSF or Bic (delta from baseline) for non-burst frequency (Hz) [% of spikes emitted in bursts, burst event frequency (Hz), burst duration (ms), and intra burst frequency (Hz) (HSD, ^∗P < 0.05 compared with all pre injection time-bins)]. Symbols are mean ± SEM for each group; lines are individual subjects. See main text for detailed statistics.)