Extracerebellar role for Cerebellin1: modulation of dendritic spine density and synapses in striatal medium spiny neurons.

Cerebellin1 (Cbln1) is a secreted glycoprotein that was originally isolated from the cerebellum and subsequently found to regulate synaptic development and stability. Cbln1 has a heterogeneous distribution in brain, but the only site in which it has been shown to have central effects is the cerebellar cortex, where loss of Cbln1 causes a reduction in granule cell-Purkinje cell synapses. Neurons of the thalamic parafascicular nucleus (PF), which provide glutamatergic projections to the striatum, also express high levels of Cbln1. We first examined Cbln1 in thalamostriatal neurons and then determined if cbln1 knockout mice exhibit structural deficits in striatal neurons. Virtually all PF neurons express Cbln1-immunoreactivity (-ir). In contrast, only rare Cbln1-ir neurons are present in the central medial complex, the other thalamic region that projects heavily to the dorsal striatum. In the striatum Cbln1-ir processes are apposed to medium spiny neuron (MSN) dendrites; ultrastructural studies revealed that Cbln1-ir axon terminals form axodendritic synapses with MSNs. Tract-tracing studies found that all PF cells retrogradely labeled from the striatum express Cbln1-ir. We then examined the dendritic structure of Golgi-impregnated MSNs in adult cbln1 knockout mice. MSN dendritic spine density was markedly increased in cbln1(-/-) mice relative to wildtype littermates, but total dendritic length was unchanged. Ultrastructural examination revealed an increase in the density of MSN axospinous synapses in cbln1(-/-) mice, with no change in postsynaptic density length. Thus, Cbln1 determines the dendritic structure of striatal MSNs, with effects distinct from those seen in the cerebellum.

Distribution of serotonin 5-HT(2A) receptors in afferents of the rat striatum.

Treatment with conventional antipsychotic drugs (APDs) is accompanied by extrapyramidal side effects (EPS), which are thought to be due to striatal dopamine D(2) receptor blockade. In contrast, treatment with atypical APDs is marked by a low incidence or absence of EPS. The reduced motor side effect liability of atypical APDs has been attributed to a high serotonin 5-HT(2A) receptor affinity coupled with a relatively low D(2) affinity. Despite the high density of 5-HT(2A) binding sites in the striatum, there are few detectable 5-HT(2A) mRNA-expressing neurons in the striatum. This suggests that most striatal 5-HT(2A) receptors are heteroceptors located on afferent axons. A combined retrograde tracer-immunohistochemistry method was used to determine the sites of origin of striatal 5-HT(2A)-like immunoreactive axons. 5-HT(2A)-like immunoreactive neurons in both the cortex and globus pallidus were retrogradely labeled from the striatum; very few nigrostriatal or thalamostriatal neurons expressed 5-HT(2A)-like immunoreactivity. Within the striatum, parvalbumin-containing interneurons displayed 5-HT(2A) immunolabeling; these neurons are the targets of cortical and pallidal projections. Our data indicate that cortico- and pallido-striatal neurons are the major source of 5-HT(2A) receptor binding in the striatum, and suggest that cortico- and pallido-striatal neurons are strategically positioned to reduce the motor side effects that accompany striatal D(2) receptor blockade or are seen in parkinsonism.

Stoichiometry of tyrosine hydroxylase phosphorylation in the nigrostriatal and mesolimbic systems in vivo: effects of acute haloperidol and related compounds.

Electrical stimulation of the medial forebrain bundle increases (32)P incorporation into striatal tyrosine hydroxylase (TH) at Ser (19), Ser(31), and Ser(40). In the present studies, the effects of acute haloperidol and related drugs on sitespecific TH phosphorylation stoichiometry (PS) in the nigrostriatal and mesolimbic systems were determined by quantitative blot immunolabeling using phosphorylation statespecific antibodies. The striatum (Str), substantia nigra (SN), nucleus accumbens (NAc), and ventral tegmental area (VTA) from Sprague-Dawley rats were harvested 30-40 min after a single injection of either vehicle, haloperidol (2 mg/kg), raclopride (2 mg/kg), clozapine (30 mg/kg), or SCH23390 (0.5 mg/kg). In vehicle-injected control rats, Ser(19) PS was 1.5- to 2. 5-fold lower in Str and NAc than in SN and VTA, Ser(31) PS was two-to fourfold higher in Str and NAc than in SN and VTA, and Ser(40) PS was similar between the terminal field and cell body regions. After haloperidol, Ser(40) PS increased twofold in Str and NAc, whereas a smaller increase in SN and VTA was observed. The effects of haloperidol on Ser(19) PS were similar to those on Ser(40) in each region; however, haloperidol treatment increased Ser(31) PS at least 1.6-fold in all regions. The effects of raclopride on TH PS were comparable to those of haloperidol, whereas clozapine treatment increased TH PS at all sites in all regions. By contrast, the effects of SCH23390 on TH PS were relatively small and restricted to the NAc. The stoichiometries of site-specific TH phosphorylation in vivo are presented for the first time. The nigrostriatal and mesolimbic systems have common features of TH PS, distinguished by differences in TH PS between the terminal field and cell body regions and by dissimilar increases in TH PS in the terminal field and cell body regions after acute haloperidol.

The distribution and origin of the calretinin-containing innervation of the nucleus accumbens of the rat.

The nucleus accumbens of the rat consists of several subregions that can be distinguished on the basis of histochemical markers. For example, the calcium-binding protein calbindin D28k is a useful marker of the core compartment of the nucleus accumbens. Calretinin, another calcium-binding protein, is found in a dense fibre plexus in the accumbal shell and septal pole regions. The source of the accumbal calretinin innervation is not known. We examined the distribution of calretinin in the nucleus accumbens and used tract-tracing and lesion methods to determine the source of this calretinin innervation. Intense calretinin immunoreactivity was present in the medial shell, but the density of calretinin axons diminished sharply in the ventrolateral shell. Regions of dense calretinin immunostaining and those areas with calbindin-like immunoreactive cell bodies were generally segregated in the nucleus accumbens, although some overlap in the transition region between the core and shell was seen. Small clusters of calretinin-immunoreactive fibres were seen in the core, where they were restricted to calbindin-negative patches. Injections of the anterograde tracer biotinylated dextran amine into the paraventricular thalamic nucleus labelled fibres in calretinin-rich regions of the accumbens. Conversely, injections of Fluoro-gold into the accumbal shell retrogradely labelled numerous cells in the paraventricular thalamic nucleus that were calretinin-immunoreactive. Electrolytic lesions of the paraventricular thalamic nucleus reduced calretinin levels in the shell by approximately 80%. These data indicate that the calretinin innervation of the nucleus accumbens is derived primarily from the thalamic paraventricular nucleus, and marks accumbal territories that are largely complementary to those defined by calbindin.

DOI-Induced activation of the cortex: dependence on 5-HT2A heteroceptors on thalamocortical glutamatergic neurons.

Administration of the hallucinogenic 5-HT(2A/2C) agonist 1-[2, 5-dimethoxy-4-iodophenyl]-2-aminopropane (DOI) induces expression of Fos protein in the cerebral cortex. To understand the mechanisms subserving this action of DOI, we examined the consequences of pharmacological and surgical manipulations on DOI-elicited Fos expression in the somatosensory cortex of the rat. DOI dose-dependently increased cortical Fos expression. Pretreatment with the selective 5-HT(2A) antagonist MDL 100,907 completely blocked DOI-elicited Fos expression, but pretreatment with the 5-HT(2C) antagonist SB 206,553 did not modify DOI-elicited Fos expression. These data suggest that DOI acts through 5-HT(2A) receptors to increase cortical Fos expression. However, we found that DOI did not induce Fos in cortical 5-HT(2A) immunoreactive neurons but did increase expression in a band of neurons spanning superficial layer V to deep III, within the apical dendritic fields of layer V 5-HT(2A)-immunoreactive cells. This band of Fos immunoreactive neurons was in register with anterogradely labeled axons from the ventrobasal thalamus, which have previously been shown to be glutamatergic and express the 5-HT(2A) transcript. The effects of DOI were markedly reduced in animals pretreated with the AMPA/KA antagonist GYKI 52466, and lesions of the ventrobasal thalamus attenuated DOI-elicited Fos expression in the cortex. These data suggest that DOI activates 5-HT(2A) receptors on thalamocortical neurons and thereby increases glutamate release, which in turn drives Fos expression in cortical neurons through an AMPA receptor-dependent mechanism. These data cast new light on the mechanisms of action of hallucinogens.

The patterns of afferent innervation of the core and shell in the "accumbens" part of the rat ventral striatum: immunohistochemical detection of retrogradely transported fluoro-gold.

Recent data have emphasized the neurochemically distinct nature of subterritories in the accumbens part of the rat ventral striatum termed the core, shell, and rostral pole. In order to gain a more comprehensive understanding of how afferents are distributed relative to these subterritories, immunohistochemical detection of retrogradely transported Fluoro-Gold was carried out following iontophoretic injections intended to involve selectively one of the subterritories. The data revealed that a number of cortical afferents of the medial shell and core originate in separate areas, i.e., the dorsal peduncular, infralimbic, and posterior piriform cortices (to medial shell) and the dorsal prelimbic, anterior agranular insular, anterior cingulate, and perirhinal cortices (to core). The lateral shell and rostral pole are innervated by cortical structures that also project either to the medial shell or core. The orbital, posterior agranular insular, and entorhinal cortices, hippocampus, and basal amygdala were observed to innervate the accumbens in a topographic manner. Following core injections, strong bilateral cortical labeling was observed. Few labeled cortical cells were observed contralaterally following injections in the medial shell. Intermediate numbers of labeled neurons were observed in contralateral cortices following lateral shell injections. Robust subcortical labeling in a variety of structures in the ventral forebrain, lateral hypothalamus, deep temporal lobe, and brainstem was observed after shell injections, particularly those that involved the caudal dorsomedial extremity of the shell, i.e., its "septal pole." Selective ipsilateral labeling of subcortical structures in the basal ganglia circuitry was observed following injections in the core and, to a lesser extent, lateral shell. It was concluded that a number of afferent systems exhibit varying degrees of segregation with respect to the accumbal subterritories.

Activation of the locus coeruleus induced by selective stimulation of the ventral tegmental area.

The effects of selective stimulation of perikarya, but not axons of passage, within the ventral tegmental area (VTA) on the locus coeruleus (LC) noradrenergic system were examined. Anterograde and combined retrograde-immunohistochemical studies indicated both dopaminergic and non-dopaminergic projections to the region of the LC originating from the VTA. Kainic acid (KA) stimulation of the VTA resulted in a dose-dependent increase in the levels of the dopamine metabolite dihydroxyphenylacetic acid (DOPAC) in the prefrontal cortex, and also elevated levels of the norepinephrine (NE) metabolite 3-methoxy-4-hydroxyphenolglycol (MHPG). Prefrontal cortical MHPG levels did not increase in response to vehicle injection or KA infusion into the hippocampus, nor did concentrations of this metabolite increase in the prefrontal cortex in response to intra-VTA KA in animals with neurotoxic lesions of the VTA. KA injection into the VTA resulted in increased MHPG levels in the hippocampus, but not the hypothalamus. Dorsal noradrenergic bundle knife cuts prevented the KA-elicited prefrontal cortical MHPG increase. These data suggest that stimulation of the mesocoeruleo dopaminergic projection arising from the VTA results in selective excitation of the LC-derived dorsal bundle noradrenergic system.

Mesencephalic dopamine modulation of pituitary and central beta-endorphin: relation to food intake regulation.

Pituitary and central beta-endorphin have been implicated in the regulation of food intake. It has been suggested that an elevation in hypophyseal beta-endorphin represents the genetic defect in the obese mutant Zucker rat. Both pituitary and central beta-endorphin systems appear to interact with dopamine. We have therefore examined hypophyseal, hypothalamic, and basal forebrain levels of beta-endorphin in the obese Zucker rat, its lean littermate, and lean littermates sustaining neurotoxic lesions of the A10 dopamine cell group in the ventral mesencephalon. The obese mutant exhibits elevated pituitary, but not central, beta-endorphin levels relative to lean littermates. A10 lesions result in a marked increase in both pituitary and hypothalamic beta-endorphin levels, and tend to decrease the amount of the peptide in the basal forebrain. These lesions do not result in either increased food intake or body weight. These data therefore suggest that elevated pituitary beta-endorphin levels do not mediate obesity in the Zucker rat, and also demonstrate that both central and pituitary beta-endorphin are modulated by a dopamine system originating in the ventral mesencephalon.