Lynn Churchill

Specificity in the projection patterns of accumbal core and shell in the rat

The efferent projections of the core and shell areas of the nucleus accumbens were studied with a combination of anterograde and retrograde tract-tracing methods, including Phaseolus vulgaris-leucoagglutinin, horseradish peroxidase and fluorescent tracers. Both the core and shell regions project to pallidal areas, i.e. ventral pallidum and entopeduncular nucleus, with a distinct topography in the sense that the core projection is located in the dorsolateral part of ventral pallidum, whereas the shell projects to the medial part of the subcommissural ventral pallidum. Both regions of the accumbens also project to mesencephalon with a bias for the core projection to innervate the substantia nigra-lateral mesencephalic tegmentum, and for the shell projection to reach primarily the ventral tegmental-paramedian tegmentum area. The most pronounced differences between core and shell projections exist in regard to the hypothalamus and extended amygdala. Whereas the core projects primarily to the entopeduncular nucleus including a part that invades the lateral hypothalamus, the shell, in addition, projects diffusely throughout the rostrocaudal extent of the lateral hypothalamus as well as to the extended amygdala, especially its sublenticular part. Both the core and shell of the accumbens have unmistakable striatal characteristics both histologically and in their connectional patterns. The shell, however, has additional features that are reminiscent of the recently described extended amygdala [Alheid G.F. and Heimer L. (1988) Neuroscience 27, 1-39; de Olmos J.S. et al. (1985) In The Rat Nervous System, pp. 223-334]; in fact, the possibility exists that the shell represents a transitional zone that seems to characterize most of the fringes of the striatal complex, where it adjoins the extended amygdala.

GABA and enkephalin projection from the nucleus accumbens and ventral pallidum to the ventral tegmental area

GABAergic and enkephalinergic afferents to the ventral tegmental area were investigated in the rat using retrograde tracing techniques combined with in situ hybridization. Following iontophoretic deposit of Fluoro-Gold in the ventral tegmental area labeling in the forebrain was most dense in the shell of the nucleus accumbens, rostral ventromedial ventral pallidum and diagonal band of Broca. A smaller density was also observed in the lateral septum. In these forebrain regions, the portion of retrogradely labeled cells that contained mRNA for glutamate decarboxylase ranged from 25% to 50%, whereas only 5% to 15% were double-labeled for preproenkephalin mRNA. Cells double-labeled with either glutamate decarboxylase or preproenkephalin mRNA were most numerous in the lateral septum, shell of the nucleus accumbens, rostral ventral pallidum and diagonal band of Broca. Large Fluoro-Gold deposits which invaded the medial substantia nigra resulted in a significant number of retrogradely labeled cells in the core of the nucleus accumbens, and a portion of these neurons also contained mRNA for glutamate decarboxylase or preproenkephalin. These data demonstrate the presence of GABAergic and enkephalinergic neurons projecting from the nucleus accumbens, ventral pallidum and diagonal band of Broca to the ventral tegmental area.

Repeated cocaine alters glutamate receptor subunit levels in the nucleus accumbens and ventral tegmental area of rats that develop behavioral sensitization.

Abstract: Increased glutamate transmission in the nucleus accumbens and ventral tegmental area has been proposed as a mechanism underlying sensitized behavioral responses to repeated cocaine administration. GluR1, GluR2/3, and NMDAR1 subunits of glutamate receptors were quantified from immunoblots in these brain nuclei in rats at 24 h and 3 weeks after discontinuing 1 week of daily cocaine injections. Motor behavior was monitored after the first and last injections of daily cocaine, and those rats that showed >20% increase in motor activity after the last compared with the first injection were considered to have developed behavioral sensitization. The subjects that developed behavioral sensitization showed a significant increase in GluR1 levels in the nucleus accumbens at 3 weeks but not at 24 h of withdrawal. Conversely, sensitized animals showed a significant increase in NMDAR1 and GluR1 levels in the ventral tegmental area at 1 day but not at 3 weeks of withdrawal. None of these increases occurred in the rats exposed to daily cocaine that did not develop behavioral sensitization (<20% increase in motor activity), and no changes were measured in the level of GluR2/3 in any treatment group. The functional importance of the increases in glutamate receptor subunit levels is suggested by the fact that the changes were present only in rats that developed behavioral sensitization to repeated cocaine administration.

Autoradiographic localization of γ-aminobutyric acidA receptors within the ventral tegmental area

Destruction of intrinsic neurons in the ventral tegmental area (VTA) with the excitotoxin, quinolinic acid produced a significant decrease (80%) in [3H]muscimol binding to GABAA receptors within the parabrachial pigmented and paranigral nuclei of the VTA. Selective destruction of the dopaminergic neurons with 6-hydroxydopamine (6-OHDA) did not reduce [3H]muscimol binding within the VTA. However, the destruction of dopaminergic neurons did produce an increase (20%) in [3H]muscimol binding contralateral to the lesion, suggesting a reduction in the GABAergic innervation to this region. Additionally, destruction of the VTA afferents with quinolinic acid injections in the medial accumbens failed to produce alterations in [3H]muscimol binding within the VTA. These results are consistent with the predominant localization of GABAA receptors to non-dopaminergic neurons intrinsic to the VTA.

Involvement of the pallidal-thalamocortical circuit in adaptive behavior.

ABSTRACT: Interconnections among the ventral mesencephalon, nucleus accumbens, and ventral pallidum are critical in the initiation of adaptive behavioral responses to environmental stimuli. Within this circuit are two highly topographically organized subcircuits that are differentially interconnected with limbic and motor circuitry in the brain. However, there is not a great deal of anatomical interconnection between the limbic and motor subcircuits. A polysynaptic connection between the two subcircuits involves projections from the limbic ventral pallidum to the mediodorsal thalamus to the prefrontal cortex back to the motor regions of the nucleus accumbens. In the present report we show that this connection is critical in the expression of motor behavior elicited by opioids and the capacity of a rat to perform in a task requiring spatial working memory.

Expression of D1 receptor mRNA in projections from the forebrain to the ventral tegmental area

In situ hybridization was combined with Fluoro‐Gold retrograde labeling to determine if cells projecting from the forebrain to the ventral tegmental area (VTA) express D1 receptor mRNA. Cell counts were made in the prefrontal cortex, shell of the nucleus accumbens, and ventral pallidum to estimate the percentage of neurons projecting to the VTA that express D1 receptor mRNA. Retrogradely labeled cells were observed in the infralimbic and prelimbic regions of the prefrontal cortex, and up to 37% of the retrogradely labeled cells expressed D1 receptor mRNA. Double‐labeled cells constituted up to 89% of retrogradely labeled neurons in the rostral shell and up to 68% in the caudal shell of the nucleus accumbens. The number of retrogradely labeled cells in the ventral pallidum that were double‐labeled ranged from 13% in the rostral to less than 10% in the caudal portions. These data provide anatomical support for a role of D1 receptors in the reciprocal innervation between the forebrain and VTA. Synapse 25:205–214, 1997.

Substance P in the ventral pallidum: Projection from the ventral striatum, and electrophysiological and behavioral cinsequences of pallidal substance P

The ventral pallidum of the basal forebrain contains a high concentration of substance P and receives a massive projection from the nucleus accumbens. The present study was designed to determine whether the accumbens serves as a source for substance P-containing fibers in the ventral pallidum and characterize the function of this tachykinin peptide within the ventral pallidum. By combining in situ hybridization for messenger RNA of the substance P prohormone, beta-preprotachykinin, with Fluoro-Gold retrograde labeling from iontophoretic deposits in the ventral pallidum, a population of substance P-containing neurons was demonstrated in the shell and core components of the nucleus accumbens and the ventromedial striatum. The function of substance P within the ventral pallidum was characterized at the level of the single neuron, and the behaving animal. Electrophysiological assessment revealed that approximately 40% of the 97 ventral pallidal neurons tested were readily excited by microiontophoretic applications of substance P or a metabolically stable agonist analog, DiMeC7 [(pGlu5, MePhe8, MeGly9)-substance P5-11]. Response characteristics were distinguished from glutamate-induced excitations by a slower onset and longer duration of action. Recording sites of tachykinin-sensitive neurons were demonstrated to be located throughout the ventral pallidum and within high densities of fibers exhibiting substance P-like immunoreactivity. When behaving rats received microinjections of DiMeC7 into this same region, the animals displayed an increase in motor activity, with a response threshold of 0.1nmol per hemisphere. These results verify the existence of a substantial substance P-containing projection from the nucleus accumbens to the ventral pallidum. The projection likely serves to excite ventral pallidal neurons for these neurons readily increased firing following local exposure to tachykinins. Furthermore, an increase in motor behavior appears to be a consequence of this neuronal response.

TUMOR NECROSIS FACTOR α: ACTIVITY DEPENDENT EXPRESSION AND PROMOTION OF CORTICAL COLUMN SLEEP IN RATS

Cortical surface evoked potentials (SEPs) are larger during sleep and characterize a sleep-like state in cortical columns. Since tumor necrosis factor alpha (TNF) may be involved in sleep regulation and is produced as a consequence of waking activity, we tested the hypothesis that direct application of TNF to the cortex will induce a sleep-like state within cortical columns and enhance SEP amplitudes. We found that microinjection of TNF onto the surface of the rat somatosensory cortex enhanced whisker stimulation-induced SEP amplitude relative to a control heat-inactivated TNF microinjection. We also determined if whisker stimulation enhanced endogenous TNF expression. TNF immunoreactivity (IR) was visualized after 2 h of deflection of a single whisker on each side. The number of TNF-IR cells increased in layers II-IV of the activated somatosensory barrel column. In two separate studies, unilateral deflection of multiple whiskers for 2 h increased the number of TNF-IR cells in layers II-V in columns that also exhibited enhanced cellular ongogene (Fos-IR). TNF-IR also colocalized with NeuN-IR suggesting that TNF expression was in neurons. Collectively these data are consistent with the hypotheses that TNF is produced in response to neural activity and in turn enhances the probability of a local sleep-like state as determined by increases in SEP amplitudes.

GHRH and IL1β increase cytoplasmic Ca2+ levels in cultured hypothalamic GABAergic neurons

GHRH and IL1beta regulate sleep via the hypothalamus. However, actions of these substances on neurons are poorly understood. In this study, we found both GHRH (100 nM) and IL1beta (1.2 pM) acutely increased cytosolic Ca(2+) in 7.6 and 4.0% of cultured hypothalamic neurons tested, respectively, and 1.2% of neurons responded to both. The neurons that responded were mostly GABAergic (96, 81, and 100% for GHRH, IL1beta, and dual-responsive neurons, respectively).

Fos-immunoreactivity in the hypothalamus: Dependency on the diurnal rhythm, sleep, gender, and estrogen

Diurnal variations and sleep deprivation-induced changes in the number of Fos-immunoreactive (Fos-IR) neurons in various hypothalamic/preoptic nuclei were studied in the rat. The nuclei implicated in sleep regulation, the ventrolateral preoptic (VLPO), median preoptic (MnPO), and suprachiasmatic (SCN, dorsomedial subdivision) nuclei, displayed maximum c-fos expression in the rest (light) period. Sleep deprivation (S.D.) suppressed Fos-IR in the dorsomedial subdivision of SCN but failed to alter Fos in the VLPO. Fos-IR increased in the VLPO during recovery after S.D. A nocturnal rise in Fos expression was detected in the arcuate (ARC), anterodorsal preoptic (ADP) and anteroventral periventricular (AVPV) nuclei whereas the lateroanterior hypothalamic nucleus (LA) and the ventrolateral subdivision of SCN did not display diurnal variations. S.D. stimulated Fos expression in the ARC, ADP, and LA. Statistically significant, albeit modest, differences were noted in the number of Fos-IR cells between males and cycling female (estrus/diestrus) in the VLPO, MnPO, ARC, LA, and AVPV, and the female ADP did not display diurnal variations. Ovariectomy (OVX) was followed by marked reduction in Fos expression in the VLPO, SCN, and AVPV, and the diurnal rhythm decreased in the VLPO, and vanished in the dorsomedial SCN, and AVP. Estrogen administration to OVX female rats stimulated Fos expression in most nuclei, and the lost diurnal variations reoccurred. In contrast, castration of male rats had little effect on Fos expression (slight rises in diurnal Fos in the ARC and ventrolateral SCN). The results suggest that Fos expression is highly estrogen-dependent in many hypothalamic nuclei including those that have been implicated in sleep regulation.