Dissociable effects of dopamine D1 and D2 receptors on compulsive ingestion and pivoting movements elicited by disinhibiting the ventral pallidum.

Previous studies have shown that infusion of a GABAA receptor antagonist, such as bicuculline (bic), into the ventral (pallidum VP) of rats elicits vigorous ingestion in sated subjects and abnormal pivoting movements. Here, we assessed if the ingestive effects generalize to the lateral preoptic area (LPO) and tested both effects for modulation by dopamine receptor signaling. Groups of rats received injections of the dopamine D2 receptor antagonist, haloperidol (hal), the D1 antagonist, SCH-23390 (SCH), or vehicle (veh) followed by infusions of bic or veh into the VP or LPO. Ingestion effects were not observed following LPO bic infusions. Compulsive ingestion associated with VP activation was attenuated by hal, but not SCH. VP bic-elicited pivoting was attenuated by neither hal, nor SCH.

The lateral preoptic area and ventral pallidum embolden behavior.

While recently completing a study of the effects of stimulating the lateral preoptic area (LPO) and ventral pallidum (VP) on locomotion and other movements, we also noticed LPO and VP effects on motivational drive and threat tolerance. Here, we have investigated these latter effects by testing conditioned place preference (CPP), behavior on the elevated plus maze (EPM) and the willingness of sated rats to occupy a harshly lit open field center to acquire sweet pellets, a measure of threat tolerance, following infusions of vehicle or bicuculline (bic) into the LPO and VP. LPO-bic infusions robustly increased total locomotion, and, in direct proportion, occupancy of both the harshly lit field center and open arms of the EPM. LPO bic also generated CPP, but did not increase sweet pellet ingestion. These effects were attenuated by dopamine D1 and D2 receptor antagonists, whether given individually or as a cocktail and systemically or infused bilaterally into the nucleus accumbens. VP-bic infusions did not increase total locomotion, but preferentially increased field center occupancy. VP-bic-infused rats compulsively ingested sweet pellets and did so even under the spotlight, whereas harsh illumination suppressed pellet ingestion in the control groups. VP bic produced CPP and increased open arm occupancy on the EPM. These effects were attenuated by pretreatment with dopamine receptor antagonists given systemically or as bilateral infusions into the VP, except for % distance in the field center (by D1 or D2 antagonists) and pellet ingestion (by D1 antagonist). Thus, boldness generated in association with LPO activation is tightly tied to locomotor activation and, as is locomotion itself, strongly DA dependent, whereas that accompanying stimulation of the VP is independent of locomotor activation and, at least in part, DA signaling. Furthermore, respective emboldened behaviors elicited from neither LPO nor VP could clearly be attributed to goal pursuit. Rather, emboldening of behavior seems more to be a fixed action response not fundamentally different than previously for reported locomotion, pivoting, backing, gnawing, and eating elicited by basal forebrain stimulation.

Lateral preoptic and ventral pallidal roles in locomotion and other movements.

The lateral preoptic area (LPO) and ventral pallidum (VP) are structurally and functionally distinct territories in the subcommissural basal forebrain. It was recently shown that unilateral infusion of the GABAA receptor antagonist, bicuculline, into the LPO strongly invigorates exploratory locomotion, whereas bicuculline infused unilaterally into the VP has a negligible locomotor effect, but when infused bilaterally, produces vigorous, abnormal pivoting and gnawing movements and compulsive ingestion. This study was done to further characterize these responses. We observed that bilateral LPO infusions of bicuculline activate exploratory locomotion only slightly more potently than unilateral infusions and that unilateral and bilateral LPO injections of the GABAA receptor agonist muscimol potently suppress basal locomotion, but only modestly inhibit locomotion invigorated by amphetamine. In contrast, unilateral infusions of muscimol into the VP affect basal and amphetamine-elicited locomotion negligibly, but bilateral VP muscimol infusions profoundly suppress both. Locomotor activation elicited from the LPO by bicuculline was inhibited modestly and profoundly by blockade of dopamine D2 and D1 receptors, respectively, but was not entirely abolished even under combined blockade of dopamine D1 and D2 receptors. That is, infusing the LPO with bic caused instances of near normal, even if sporadic, invigoration of locomotion in the presence of saturating dopamine receptor blockade, indicating that LPO can stimulate locomotion in the absence of dopamine signaling. Pivoting following bilateral VP bicuculline infusions was unaffected by dopamine D2 receptor blockade, but was completely suppressed by D1 receptor blockade. The present results are discussed in a context of neuroanatomical and functional organization underlying exploratory locomotion and adaptive movements.

Abundant collateralization of temporal lobe projections to the accumbens, bed nucleus of stria terminalis, central amygdala and lateral septum.

Behavioral flexibility is subserved in part by outputs from the cerebral cortex to telencephalic subcortical structures. In our earlier evaluation of the organization of the cortical-subcortical output system (Reynolds and Zahm, J Neurosci 25:11757-11767, 2005), retrograde double-labeling was evaluated in the prefrontal cortex following tracer injections into pairs of the following subcortical telencephalic structures: caudate-putamen, core and shell of the accumbens (Acb), bed nucleus of stria terminalis (BST) and central nucleus of the amygdala (CeA). The present study was done to assess patterns of retrograde labeling in the temporal lobe after similar paired tracer injections into most of the same telencephalic structures plus the lateral septum (LS). In contrast to the modest double-labeling observed in the prefrontal cortex in the previous study, up to 60-80 % of neurons in the basal and accessory basal amygdaloid nuclei and amygdalopiriform transition area exhibited double-labeling in the present study. The most abundant double-labeling was generated by paired injections into structures affiliated with the extended amygdala, including the CeA, BST and Acb shell. Injections pairing the Acb core with the BST or CeA produced significantly fewer double-labeled neurons. The ventral subiculum exhibited modest amounts of double-labeling associated with paired injections into the Acb, BST, CeA and LS. The results raise the issue of how an extraordinarily collateralized output from the temporal lobe may contribute to behavioral flexibility.

Protracted maturation of forebrain afferent connections of the ventral tegmental area in the rat.

The mesocorticolimbic dopamine system has long attracted the interest of researchers concerned with the unique gamut of behavioral and mental health vulnerabilities associated with adolescence. Accordingly, the development of the mesocorticolimbic system has been studied extensively, but almost exclusively with regard to dopaminergic output, particularly in the nucleus accumbens and medial prefrontal cortex. To the contrary, the ontogeny of inputs to the ventral tegmental area (VTA), the source of mesocorticolimbic dopamine, has been neglected. This is not a trivial oversight, as the activity of VTA neurons, which reflects their capacity to transmit information about salient events, is sensitively modulated by inputs. Here, we assessed the development of VTA afferent connections using the ß subunit of cholera toxin (Ctß) as a retrograde axonal tracer in adolescent (postnatal day 39) and early adult (8-9-week-old) rats. After intra-VTA injections of Ctß, adolescent and early adult animals exhibited qualitatively similar distributions of retrogradely labeled neurons in the sense that VTA-projecting neurons were present at all of the same rostrocaudal levels in all of the same structures in both age groups. However, quantitation of retrogradely labeled neurons revealed that adolescent brains, compared with early adult brains, had significantly fewer VTA-projecting neurons preferentially within an interconnected network of cortical and striatopallidal forebrain structures. These findings provide a novel perspective on the development of the mesocorticolimbic dopamine system and may have important implications for age-dependent specificity in the function of this system, particularly with regard to adolescent impulsivity and mental health vulnerabilities.

Oxidative stress disruption of receptor-mediated calcium signaling mechanisms.

BACKGROUND: Oxidative stress increases the cytosolic content of calcium in the cytoplasm through a combination of effects on calcium pumps, exchangers, channels and binding proteins. In this study, oxidative stress was produced by exposure to tert-butyl hydroperoxide (tBHP); cell viability was assessed using a dye reduction assay; receptor binding was characterized using [3H]N-methylscopolamine ([3H]MS); and cytosolic and luminal endoplasmic reticulum (ER) calcium concentrations ([Ca2+]i and [Ca2+]L, respectively) were measured by fluorescent imaging. RESULTS: Activation of M3 muscarinic receptors induced a biphasic increase in [Ca2+]i: an initial, inositol trisphosphate (IP3)-mediated release of Ca2+ from endoplasmic reticulum (ER) stores followed by a sustained phase of Ca2+ entry (i.e., store-operated calcium entry; SOCE). Under non-cytotoxic conditions, tBHP increased resting [Ca2+]i; a 90 minute exposure to tBHP (0.5-10 mM ) increased [Ca2+]i from 26 to up to 127 nM and decreased [Ca2+]L by 55%. The initial response to 10 µM carbamylcholine was depressed by tBHP in the absence, but not the presence, of extracellular calcium. SOCE, however, was depressed in both the presence and absence of extracellular calcium. Acute exposure to tBHP did not block calcium influx through open SOCE channels. Activation of SOCE following thapsigargin-induced depletion of ER calcium was depressed by tBHP exposure. In calcium-free media, tBHP depressed both SOCE and the extent of thapsigargin-induced release of Ca2+ from the ER. M3 receptor binding parameters (ligand affinity, guanine nucleotide sensitivity, allosteric modulation) were not affected by exposure to tBHP. CONCLUSIONS: Oxidative stress induced by tBHP affected several aspects of M3 receptor signaling pathway in CHO cells, including resting [Ca2+]i, [Ca2+]L, IP3 receptor mediated release of calcium from the ER, and calcium entry through the SOCE. tBHP had little effect on M3 receptor binding or G protein coupling. Thus, oxidative stress affects multiple aspects of calcium homeostasis and calcium dependent signaling.

Honokiol blocks store operated calcium entry in CHO cells expressing the M3 muscarinic receptor: honokiol and muscarinic signaling.

BACKGROUND: Honokiol, a cell-permeable phenolic compound derived from the bark of magnolia trees and present in Asian herbal teas, has a unique array of pharmacological actions, including the inhibition of multiple autonomic responses. We determined the effects of honokiol on calcium signaling underlying transmission mediated by human M3 muscarinic receptors expressed in Chinese hamster ovary (CHO) cells. Receptor binding was determined in radiolabelled ligand binding assays; changes in intracellular calcium concentrations were determined using a fura-2 ratiometric imaging protocol; cytotoxicity was determined using a dye reduction assay. RESULTS: Honokiol had a potent (EC50 ≈ 5 µmol/l) inhibitory effect on store operated calcium entry (SOCE) that was induced by activation of the M3 receptors. This effect was specific, rapid and partially reversible, and was seen at concentrations not associated with cytotoxicity, inhibition of IP3 receptor-mediated calcium release, depletion of ER calcium stores, or disruption of M3 receptor binding. CONCLUSIONS: It is likely that an inhibition of SOCE contributes to honokiol disruption of parasympathetic motor functions, as well as many of its beneficial pharmacological properties.