Locus coeruleus neuronal activity determines proclivity to consume alcohol in a selectively-bred line of rats that readily consumes alcohol

Abstract

Sprague-Dawley rats selectively-bred for susceptibility to stress in our laboratory (Susceptible, or SUS rats) voluntarily consume large amounts of alcohol, and amounts that have, as shown here, pharmacological effects, which normal rats will not do. In this paper, we explore neural events in the brain that underlie this propensity to readily consume alcohol. Activity of locus coeruleus neurons (LC), the major noradrenergic cell body concentration in the brain, influences firing of ventral tegmentum dopaminergic cell bodies of the mesocorticolimbic system (VTA-DA neurons), which mediate rewarding aspects of alcohol. We tested the hypothesis that in SUS rats alcohol potently suppresses LC activity to markedly diminish LC-mediated inhibition of VTA-DA neurons, which permits alcohol to greatly increase VTA-DA activity and rewarding aspects of alcohol. Electrophysiological single-unit recording of LC and VTA-DA activity showed that in SUS rats alcohol decreased LC burst firing much more than in normal rats and as a result markedly increased VTA-DA activity in SUS rats while having no such effect in normal rats. Consistent with this, in a behavioral test for reward using conditioned place preference (CPP), SUS rats showed alcohol, given by intraperitoneal (i.p.) injection, to be rewarding. Next, manipulation of LC activity by microinfusion of drugs into the LC region of SUS rats showed that (a) decreasing LC activity increased alcohol intake and increasing LC activity decreased alcohol intake in accord with the formulation described above, and (b) increasing LC activity blocked both the rewarding effect of alcohol in the CPP test and the usual alcohol-induced increase in VTA-DA single-unit activity seen in SUS rats. An important ancillary finding in the CPP test was that an increase in LC activity was rewarding by itself, while a decrease in LC activity was aversive; consequently, effects of LC manipulations on alcohol-related reward in the CPP test were perhaps even larger than evident in the test. Finally, when increased LC activity was associated with (i.e., conditioned to) i.p. alcohol, subsequent alcohol consumption by SUS rats was markedly reduced, indicating that SUS rats consume large amounts of alcohol because of rewarding physiological consequences requiring increased VTA-DA activity. The findings reported here are consistent with the view that the influence of alcohol on LC activity leading to changes in VTA-DA activity strongly affects alcohol-mediated reward, and may well be the basis of the proclivity of SUS rats to avidly consume alcohol.

Keywords: Alcohol; Conditioned place preference; Dopamine; Locus coeruleus; Selectively-bred rats; Ventral tegmentum.

Fig. 1

Firing activity of locus coeruleus (LC) neurons in SUS and randomly-bred normal SD rats in response to alcohol injected intraperitoneally (i.p.). Single-unit electrophysiological activity (spikes per second [Hz]) of LC neurons (means and standard errors) is shown. Following determination of baseline activity (0.0 dose), the effect of increasing (cumulative) doses of i.p.-injected alcohol was assessed on Spontaneous firing rate (Left) and Sensory-evoked burst firing (Right). Number of rats/group = 6, one unit per rat.

Fig. 2

Firing activity of dopaminergic neurons in the ventral tegmental area (VTA-DA neurons) in SUS and randomly-bred normal SD rats in response to alcohol injected intraperitoneally (i.p.). Single-unit electrophysiological activity (spikes per second [Hz]) of VTA-DA neurons (means and standard errors) is shown. Following determination of baseline activity (0.0 dose), the effect of increasing (cumulative) doses of i.p.-injected alcohol was assessed on Spontaneous firing rate (Left) and percentage of spikes occurring within bursts (Right). Number of rats/group = 6; one unit per rat.

Fig. 3

Consumption of 10% ethanol solution (g/kg) in 1.0 h by SUS rats after no procedure (Pre-test) or after infusion of artificial CSF vehicle (Veh) and drugs to either decrease LC activity (clonidine [Clon]) or increase LC activity (substance P + Idazoxan [Sub P + Ida]). Means and standard errors are shown. The same rats (n = 7) underwent all phases of this study. In the week following the pre-test measure, all animals received two infusions, one of vehicle (Veh 1) and the other of Clon to decrease LC activity, with the infusions given 4 days apart (with consumption test after each infusion); three rats received vehicle first and then Clon and four rats received infusions in the reverse order. In the following week, all animals again received two infusions, one of vehicle (Veh 2) and the other of Sub P + Ida to increase LC activity, with infusions again 4 days apart; again, three rats received vehicle first and then drug while four rats received infusions in reverse order. * = Clon differs from Veh 1, p < .005; ** = Sub P + Ida differs from Veh 2, p < .003.

Fig. 4

A (left). Conditioned Place Preference (CPP) of SUS rats produced by injections of alcohol (1.0 g/kg) versus saline shown by time (seconds) spent during a 30-min test session on either the alcohol-paired side or the saline-paired side of the CPP test chamber. Means and standard errors are shown. n = 6. * = significant difference (p < 0.002) in time spent on alcohol-associated side of CPP apparatus before vs. after conditioning. B (right). Blood Alcohol Concentration (BAC) found in SUS rats after voluntary consumption of a 10% alcohol solution and after injection of 1.0 g/kg alcohol as used in CPP conditioning. Blood samples were taken 45 min after onset of ethanol presentation for drinking and after i.p. injection. Means and standard errors are shown. n = 6 (same rats as in A).

Fig. 5

A (Left). Conditioned Place Preference (CPP) of SUS rats produced by i.p. injection of alcohol (1.0 g/kg) versus saline shown by animals that had been infused into locus coeruleus (LC) with vehicle (aCSF) (n = 6) or Substance P + Idazoxan (Sub P + Ida) to increase LC activity (n = 6) or clonidine (Clon) to decrease LC activity (n = 7) immediately before the injections of alcohol in this procedure. Shown is time spent during the 30-min test session on the alcohol-paired side of two-compartment CPP apparatus; at top is shown time (in seconds) spent on the alcohol-paired side in the test session, and at bottom is shown the percentage of total time in CPP apparatus during the test session spent on the alcohol-paired side. Means and standard errors are shown. “Baseline” shows time spent on what became the alcohol-paired side of the CPP apparatus before any injection of alcohol or any other procedure; “Test day” shows time spent on that side after i.p. injections of alcohol followed by confinement on that distinctive side of the CPP apparatus and after an equal number of injections of i.p. physiological saline (0.85% saline) and confinement on the other distinctive side of the CPP apparatus (i.e., post conditioning). Statistics: * = significantly higher than the “Baseline” for these same animals (atleast p < .02; by paired t tests), and also significantly higher than Sub P + Ida-infused rats on “Test day” (at least p < .005; by independent groups t tests). B (Right). Conditioned Place Preference (CPP) of SUS rats with details exactly as above except that all i.p. injections were physiological saline (i.e., no alcohol given). Thus, animals were infused into locus coeruleus (LC) with Sub P + Ida to increase LC activity (n = 6) or Clon to decrease LC activity (n = 7) followed by i.p. saline, and consequently this CPP procedure determined preference for, or aversion of, the side of the CPP apparatus paired with LC activity having been either increased (vs. no change in LC activity by aCSF infusion) or decreased (vs. no change in LC activity by aCSF infusion). Statistics: * = significantly different (higher or lower) than the “Baseline” for these same animals (at least p < .05 by paired t tests).

Fig. 6

Firing activity of dopaminergic neurons in the ventral tegmental area (VTA-DA neurons) in response to alcohol injected intraperitoneally (i.p.) in SUS rats infused into LC with Substance P + Idazoxan (Sub P + Ida) to increase LC activity (n = 6) or aCSF to have no effect on LC activity (n = 9). Single-unit electrophysiological activity (spikes per second [Hz]) of VTA-DA neurons (means and standard errors) is shown. Recording was done in the VTA on the side of brain ipsilateral to where unilateral infusion into LC was done. Following infusion of aCSF or drug, baseline activity was determined (0.0 dose), and then the effect of increasing (cumulative) doses of i.p.-injected alcohol was assessed on Spontaneous firing rate (Left) and percentage of spikes occurring within bursts (Right). Statistics: Analysis of both spontaneous firing rate and burst firing was done by 2-way (Group × alcohol dose) repeated-measures analysis of covariance (repeated measure for each subject across alcohol doses; covariate was the first measure after infusion shown at 0.0 alcohol dose). For spontaneous firing rate, Group factor (i.e., aCSF vs. Sub P + Ida) was significantly different (F[1,12] = 5.92, p < .032) as was the interaction (Group × alcohol dose) (F[3,36] = 2.93, p < .047). For burst firing, the Group factor was also significantly different (F[1,12] = 10.59, p < .007) as was the interaction (Group × alcohol dose) (F[3,36] = 3.84, p < .017). The alcohol dose factor was also significant (F[3,36] = 1.77, p < .043) in analysis of burst firing.

Fig. 7

Consumption of 10% alcohol solution (g/kg) in 1.0 h by SUS rats prior to their undergoing the Conditioned Place Preference (CPP) procedure (Pre CPP) and on four consumption tests at 2-week intervals after these animals had undergone the CPP conditioning procedure (Tests after CPP). Means and standard errors are shown. During the CPP procedure, all animals had received bilateral infusions into LC immediately before alcohol was injected i.p. (1.0 g/kg), some animals receiving artificial CSF to have no effect on LC activity (aCSF), other animals receiving Substance P + Idazoxan to increase LC activity (Sub P + Ida), and other animals receiving Clonidine to decrease LC activity (Clon). Statistics: ** = significantly lower (p < .02) and * = significantly lower (p < .05) than aCSF-infused animals on that test by contrasts on each test day.