Habenulo-interpeduncular Pathway Research Articles

KCTD8 and KCTD12 Facilitate Axonal Expression of GABAB Receptors in Habenula Cholinergic Neurons.

GABAB receptors in habenula cholinergic neurons mediate strong presynaptic excitation and control aversive memory expression. K+ channel tetramerization domain (KCTD) proteins are key interacting partners of GABAB receptors; it remains unclear whether and how KCTDs contribute to GABAB excitatory signaling. Here, we show that KCTD8 and KCTD12 in these neurons facilitate the GABAB receptors expression in axonal terminals and contribute to presynaptic excitation by GABAB receptors. Genetically knocking out KCTD8/12/16 or KCTD8/12, but not other combinations of the three KCTD isoforms, substantially reduced GABAB receptors-mediated potentiation of glutamate release and presynaptic Ca2+ entry in response to axonal stimulation, whereas they had no effect on GABAB-mediated inhibition in the somata of cholinergic neurons within the habenulo-interpeduncular pathway in mice of either sex. The physiological phenotypes were associated with a significant decrease in the GABAB expression within the axonal terminals but not the somata. Overexpressing either KCTD8 or KCTD12 in the KCTD8/12/16 triple knock-out mice reversed the changes in axonal GABAB expression and presynaptic excitation. In mice lacking the KCTDs, aversion-predicting cues produced stronger neuronal activation in the interpeduncular nucleus, and the infusion of GABAB agonist in this nucleus produced a weaker effect on fear extinction. Collectively, our results reveal isoform-specific roles of KCTD proteins in enriching the axonal expression of GABAB receptors, facilitating their presynaptic signaling, and modulating aversion-related memory processes.SIGNIFICANCE STATEMENT GABAB receptors represent the principal inhibitory neurotransmitter receptor, but they mediate strong presynaptic excitation in the habenulo-interpeduncular pathway and modulate aversion memory expression. KCTD proteins are integral constituents of GABAB receptors. By analyzing the physiological, neuroanatomical, and behavioral phenotypes of multiple KCTD knock-out mouse lines, we show that KCTD8 and KCTD12 facilitate the axonal expression and hence presynaptic excitation of GABAB receptors in habenula cholinergic neurons and control cued-aversion memory formation and expression in the habenulo-interpeduncular pathway. These results expand the physiological and behavioral functions of KCTDs in modulating the brain neural circuits.

Trans-inhibition of axon terminals underlies competition in the habenulo-interpeduncular pathway

Survival of animals is dependent on the correct selection of an appropriate behavioral response to competing external stimuli. Theoretical models have been proposed and underlying mechanisms are emerging to explain how one circuit is selected among competing neural circuits. The evolutionarily conserved forebrain to midbrain habenulo-interpeduncular nucleus (Hb-IPN) pathway consists of cholinergic and non-cholinergic neurons, which mediate different aversive behaviors. Simultaneous calcium imaging of neuronal cell bodies and of the population dynamics of their axon terminals reveals that signals in the cell bodies are not reflective of terminal activity. We find that axon terminals of cholinergic and non-cholinergic habenular neurons exhibit stereotypic patterns of spontaneous activity that are negatively correlated and localize to discrete subregions of the target IPN. Patch-clamp recordings show that calcium bursts in cholinergic terminals at the ventral IPN trigger excitatory currents in IPN neurons, which precede inhibition of non-cholinergic terminals at the adjacent dorsal IPN. Inhibition is mediated through presynaptic GABAB receptors activated in non-cholinergic habenular neurons upon GABA release from the target IPN. Together, the results reveal a hardwired mode of competition at the terminals of two excitatory neuronal populations, providing a physiological framework to explore the relationship between different aversive responses.

Nicotinic Receptors Underlying Nicotine Dependence: Evidence from Transgenic Mouse Models.

Nicotine underlies the reinforcing properties of tobacco cigarettes and e-cigarettes. After inhalation and absorption, nicotine binds to various nicotinic acetylcholine receptor (nAChR) subtypes localized on the pre- and postsynaptic membranes of cells, which subsequently leads to the modulation of cellular function and neurotransmitter signaling. In this chapter, we begin by briefly reviewing the current understanding of nicotine's actions on nAChRs and highlight considerations regarding nAChR subtype localization and pharmacodynamics. Thereafter, we discuss the seminal discoveries derived from genetically modified mouse models, which have greatly contributed to our understanding of nicotine's effects on the reward-related mesolimbic pathway and the aversion-related habenulo-interpeduncular pathway. Thereafter, emerging areas of research focusing on modulation of nAChR expression and/or function are considered. Taken together, these discoveries have provided a foundational understanding of various genetic, neurobiological, and behavioral factors underlying the motivation to use nicotine and related dependence processes, which are thereby advancing drug discovery efforts to promote long-term abstinence.

Left Habenular Activity Attenuates Fear Responses in Larval Zebrafish

Fear responses are defensive states that ensure survival of an organism in the presence of a threat. Perception of an aversive cue causes changes inbehavior and physiology, such as freezing and elevated cortisol, followed by a return to the baseline state when the threat is evaded [1]. Neural systems that elicit fear behaviors include the amygdala, hippocampus, and medial prefrontal cortex. However, aside from a few examples, little is known about brain regions that promote recovery from an aversive event [2]. Previous studies had implicated the dorsal habenular nuclei in regulating fear responses and boldness in zebrafish [3-7]. We now show, through perturbation of its inherent left-right (L-R) asymmetry at larval stages, that the dorsal habenulo-interpeduncular (dHb-IPN) pathway expedites the return of locomotor activity following an unexpected negativestimulus, electric shock. Severing habenular efferents to the IPN, or only those from the left dHb,prolongs the freezing behavior that follows shock.Individuals with a symmetric, right-isomerized dHb also exhibit increased freezing. In contrast, larvae that have a symmetric, left-isomerized dHb, or inwhich just the left dHb-IPN projection is optogenetically activated, rapidly resume swimming post shock. Invivo calcium imaging reveals a neuronal subset, predominantly in the left dHb, whose activation is correlated with resumption of swimming. The results demonstrate functional specialization of the left dHb-IPN pathway in attenuating the response to fear.

Role of β4* Nicotinic Acetylcholine Receptors in the Habenulo-Interpeduncular Pathway in Nicotine Reinforcement in Mice.

Nicotine exerts its psychopharmacological effects by activating the nicotinic acetylcholine receptor (nAChR), composed of alpha and/or beta subunits, giving rise to a diverse population of receptors with a distinct pharmacology. β4-containing (β4*) nAChRs are located almost exclusively in the habenulo-interpeduncular pathway. We examined the role of β4* nAChRs in the medial habenula (MHb) and the interpeduncular nucleus (IPN) in nicotine reinforcement using behavioral, electrophysiological, and molecular techniques in transgenic mice. Nicotine intravenous self-administration (IVSA) was lower in constitutive β4 knockout (KO) mice at all doses tested (7.5, 15, 30, and 60 μg/kg/infusion) compared with wild-type (WT) mice. In vivo microdialysis showed that β4KO mice have higher extracellular dopamine (DA) levels in the nucleus accumbens than in WT mice, and exhibit a differential sensitivity to nicotine-induced DA outflow. Furthermore, electrophysiological recordings in the ventral tegmental area (VTA) demonstrated that DA neurons of β4KO mice are more sensitive to lower doses of nicotine than that of WT mice. Re-expression of β4* nAChRs in IPN neurons fully restored nicotine IVSA, and attenuated the increased sensitivity of VTA DA neurons to nicotine. These findings suggest that β4* nAChRs in the IPN have a role in maintaining nicotine IVSA.

Psychiatric disorder‐related abnormal behavior and habenulointerpeduncular pathway defects in Wnt1‐cre and Wnt1‐GAL4 double transgenic mice

The neural crest is a unique structure in vertebrates. Wnt1-cre and Wnt1-GAL4 double transgenic (dTg) mice have been used in a variety of studies concerning neural crest cell lineages in which the Cre/loxP or GAL4/UAS system was applied. Here, we show psychiatric disorder-related behavioral abnormalities and histologic alterations in a neural crest-derived brain region in dTg mice. The dTg mice exhibited increased locomotor activity, decreased social interaction, and impaired short-term spatial memory and nesting behavior. The choline acetyltransferase- and vesicular glutamate transporter 2-immunoreactive habenulointerpeduncular fiber tracts that project from the medial habenular nucleus of the epithalamus to the interpeduncular nucleus of the midbrain tegmentum appeared irregular in the dTg mice. Both the medial habenula nucleus and the interpeduncular nucleus were confirmed to be derived from the neural crest. The findings of this study suggest that neural crest-derived cells have pathogenic roles in the development of psychiatric disorders and that the dTg mouse could be a useful animal model for studying the pathophysiology of mental illness such as autism and schizophrenia. Scientists that use the dTg mice as a cre-transgenic deleter line should be cautious in its possible toxicity, especially if behavioral analyses are to be performed.

Habenula “Cholinergic” Neurons Corelease Glutamate and Acetylcholine and Activate Postsynaptic Neurons via Distinct Transmission Modes

Acetylcholine is an important neurotransmitter, and the habenulo-interpeduncular projection is a major cholinergic pathway in the brain. To study the physiological properties of cholinergic transmission in the interpeduncular nucleus (IPN), we used a transgenic mouse line in which the light-gated cation channel ChannelRhodopsin-2 is selectively expressed in cholinergic neurons. Cholinergic axonal terminals were activated by light pulses, and postsynaptic responses were recorded from IPN neurons. Surprisingly, brief photostimulation produces fast excitatory postsynaptic currents that are mediated by ionotropic glutamate receptors, suggesting wired transmission of glutamate. By contrast, tetanic photostimulation generates slow inward currents that are largely mediated by nicotinic acetylcholine receptors, suggesting volume transmission of acetylcholine. Finally, vesicular transporters for glutamate and acetylcholine are coexpressed on the same axonal terminals in the IPN. These results strongly suggest that adult brain "cholinergic" neurons can corelease glutamate and acetylcholine, but these two neurotransmitters activate postsynaptic neurons via different transmission modes.

Habenular α5 nicotinic receptor subunit signalling controls nicotine intake.

Genetic variation in CHRNA5, the gene encoding the α5 nicotinic acetylcholine receptor (nAChR) subunit, increases vulnerability to tobacco addiction and lung cancer, but underlying mechanisms are unknown. Here, we report dramatically increased nicotine consumption in mice with null mutation in Chrna5. This effect was `rescued' in knockout mice by re-expressing α5 subunits in medial habenula (MHb), and recapitulated in rats through α5 subunit knockdown in MHb. Remarkably, α5 subunit knockdown in MHb did not alter the rewarding effects of nicotine but abolished the inhibitory effects of higher nicotine doses on brain reward systems. The MHb extends projections almost exclusively to the interpeduncular nucleus (IPN). We found diminished IPN activation in response to nicotine in α5 knockout mice and disruption of IPN signaling increased nicotine intake in rats. Our findings suggest that nicotine activates the habenulo-interpeduncular pathway through α5-containing nAChRs, triggering an inhibitory motivational signal that acts to limit nicotine intake.

Brain regions mediating α3β4 nicotinic antagonist effects of 18-MC on methamphetamine and sucrose self-administration

The novel iboga alkaloid congener 18-methoxycoronaridine (18-MC) is a putative anti-addictive agent that has been shown, in rats, to decrease the self-administration of several drugs of abuse. Previous work has established that 18-MC is a potent antagonist at α3β4 nicotinic receptors. Because high densities of α3β4 nicotinic receptors occur in the medial habenula and the interpeduncular nucleus and moderate densities occur in the dorsolateral tegmentum, ventral tegmental area, and basolateral amygdala, the present study was conducted to determine if 18-MC could act in these brain areas to modulate methamphetamine self-administration in rats. Local administration of 18-MC into either the medial habenula, the interpeduncular area or the basolateral amygdala decreased methamphetamine self-administration. Similar results were produced by local administration into the same brain areas of two other α3β4 nicotinic antagonists, mecamylamine and α-conotoxin AuIB. Local administration of 18-MC, or the other antagonists, into the dorsolateral tegmentum or the ventral tegmental area had no effect on methamphetamine self-administration. In contrast, local administration of 18-MC and the other antagonists decreased sucrose self-administration when administered into the dorsolateral tegmentum or basolateral amygdala but had no effect when infused into the medial habenula, interpeduncular nucleus, or ventral tegmental area. These data are consistent with the hypothesis that 18-MC decreases methamphetamine self-administration by indirectly modulating the dopaminergic mesolimbic pathway via blockade of α3β4 nicotinic receptors in the habenulo-interpeduncular pathway and the basolateral amygdala. The data also suggest that the basolateral amygdala along with a different pathway involving α3β4 receptors in the dorsolateral tegmentum mediate the effect of 18-MC on sucrose self-administration.

Effects of nicotine, methamphetamine and cocaine on extracellular levels of acetylcholine in the interpeduncular nucleus of rats

There is increasing evidence that the cholinergic habenulo-interpeduncular pathway and the dopaminergic mesolimbic pathway may jointly mediate the reinforcing properties of addictive drugs. However, the effects of addictive drug on the functioning of the habenulo-interpeduncular pathway have not been well-characterized. Thus, several drugs of abuse (i.e., nicotine, cocaine, amphetamine) have been shown to alter the morphology of the habenulo-interpeduncular pathway, causing selective degeneration of the cholinergic neurons in this area. On the other hand, morphine was shown to alter the neurochemistry of the habenulo-interpeduncular pathway, inducing biphasic changes in acetylcholine release in the interpeduncular nucleus. In order to determine the effects of cocaine, amphetamine and nicotine on cholinergic neurotransmission in the habenulo-interpeduncular pathway, levels of acetylcholine were assessed during microdialysis in freely moving rats. Nicotine (0.1 and 0.4 mg/kg s.c.) produced a dose-dependent decrease in extracellular levels of acetylcholine, while methamphetamine (1 and 4 mg/kg i.p.) produced an increase in acetylcholine release in the interpeduncular nucleus. Cocaine (5 and 20 mg/kg i.p.) produced a biphasic effect on extracellular acetylcholine release, i.e., a low dose enhanced the release of acetylcholine and a high dose decreased its release. These results suggest that the habenulo-intepeduncular pathway may be a common target for drugs of abuse and, by modulating the mesolimbic pathway, may mediate unique aspects of the rewarding effects of different drugs.

Morphine-induced changes in acetylcholine release in the interpeduncular nucleus and relationship to changes in motor behavior in rats

Owing to multiple anatomical connections and functional interactions between the habenulo-interpeduncular and the mesolimbic pathways, it has been proposed that these systems could together mediate the reinforcing properties of addictive drugs. 18-Methoxycoronaridine, an agent that reduces morphine self-administration and attenuates dopamine sensitization in the nucleus accumbens in response to repeated morphine, has been shown to produce these effects by acting in the medial habenula and interpeduncular nucleus. Acetylcholine, one of the predominant neurotransmitters in the interpeduncular nucleus, may be a major determinant of these interactions. To determine if and how morphine acts in the interpeduncular nucleus, the effects of acute and repeated administration of morphine on extracellular acetylcholine levels in this brain area were assessed. In addition, the motor behavior of rats receiving repeated morphine administration was monitored during microdialysis sessions. Acutely, morphine produced a biphasic effect on extracellular acetylcholine levels in the interpeduncular nucleus such that low and high doses of morphine (i.e., 5 and 20 mg/kg i.p.) significantly increased and decreased acetylcholine levels, respectively. Repeated administration of the same doses of morphine resulted in tolerance to the inhibitory but not to the stimulatory effects; tolerance was accompanied by sensitization to morphine-induced changes in locomotor activity and stereotypic behavior. The latter results suggest that tolerance to morphine's effect on the cholinergic habenulo-interpeduncular pathway is related to its sensitizing effects on the mesostriatal dopaminergic pathways.

18‐MC acts in the medial habenula and interpeduncular nucleus to attenuate dopamine sensitization to morphine in the nucleus accumbens

18-Methoxycoronaridine (18-MC), a novel iboga alkaloid congener, is a potential treatment for drug addiction. 18-MC has been shown to decrease self-administration of drugs (e.g., morphine, methamphetamine, nicotine) and attenuate opioid withdrawal in rats. In previous studies, systemic pretreatment with 18-MC abolished the sensitized increase in accumbens dopamine levels induced by chronic morphine administration. In vitro studies have shown that 18-MC is a potent antagonist of alpha3beta4 nicotinic receptors, and alpha3beta4 antagonism is believed to be the primary mechanism responsible for 18-MC's effects on drug self-administration and possibly on morphine-induced changes in mesolimbic dopamine. While there are very low densities of alpha3beta4 nicotinic receptors in the mesolimbic pathway, these receptors are prominently localized in the medial habenula (MHb) and in the interpeduncular nucleus (IPN). These nuclei and the habenulo-interpeduncular pathway connecting them are believed to function as part of an alternate reward pathway modulating the dopaminergic mesolimbic pathway known to be involved in drug addiction. In the present study, to determine if 18-MC acts in the MHb or in the IPN, the effects of local infusion of 18-MC into these brain areas were assessed on mesolimbic dopamine responses to acute and repeated morphine treatment. Administration of 18-MC (10 mug) into either the IPN or MHb blocked the sensitized dopamine response to repeated morphine in the nucleus accumbens; 18-MC had no effect on the dopamine response to acute morphine. The results suggest that 18-MC acts in the habenulo-interpeduncular pathway to modulate the effects of repeated morphine in the dopaminergic mesolimbic system.

Distinct structure and activity of monoamine oxidase in the brain of zebrafish (Danio rerio)

Monoamine oxidase (MAO) is a mitochondrial flavoprotein involved in the metabolism of, e.g., aminergic neurotransmitters and the parkinsonism-inducing neurotoxin 1-methyl-4-phenyl-1,2,5,6-tetrahydropyridine (MPTP). We have reported earlier MPTP-related alterations of brain catecholaminergic system in zebrafish (Danio rerio) brain. Here we describe the structural and functional properties of zebrafish MAO and the distribution of MAO mRNA and activity in zebrafish brain. The gene is located in chromosome 9 and consists of 15 exons. The amino acid composition of the active center resembles both human MAO-A and MAO-B. The enzyme displayed the highest substrate specificity for tyramine, followed by serotonin, phenylethylamine, MPTP, and dopamine; isoform-specific antagonists blocked the activity of the enzyme with equal potency. Zebrafish MAO mRNA, which was present in several tissues, and enzyme displayed differential distribution in the brain; dopaminergic cell clusters had low to moderate levels of MAO activity, whereas the highest levels of MAO activity were detected in noradrenergic and serotonergic cell groups and the habenulointerpeduncular pathway, including its caudal projection to the medial ventral rhombencephalon. The results of this study confirm the presence of functionally active MAO in zebrafish brain and other tissues and characterize the neural systems that express MAO and areas of intense activity in the brain. They also suggest that MPTP toxicity not related to MAO may affect the zebrafish brain.

18-Methoxycoronaridine acts in the medial habenula and/or interpeduncular nucleus to decrease morphine self-administration in rats

The novel iboga alkaloid congener 18-methoxycoronaridine (18-MC) is a putative anti-addictive agent that has been shown, in rats, to decrease the self-administration of morphine and other drugs of abuse. Previous work has established that 18-MC is a potent antagonist at α3β4 nicotinic receptors. Because α3β4 nicotinic receptors in the brain are preferentially located in the medial habenula and the interpeduncular nucleus, the present study was conducted to determine if 18-MC could act in these brain areas to modulate morphine self-administration in rats. Local administration of 18-MC into either the medial habenula or the interpeduncular area decreased morphine self-administration while having no effect on responding for a non-drug reinforcer (sucrose). Similar results were produced by local administration into the same brain areas of two other α3β4 nicotinic antagonists, mecamylamine and α-conotoxin AuIB. Local administration of 18-MC into the ventral tegmental area had no effect on morphine self-administration. These and other data are consistent with the hypothesis that 18-MC decreases morphine self-administration by blocking α3β4 nicotinic receptors in the habenulo-interpeduncular pathway.

Mapping of the habenulo-interpeduncular pathway in living mice using manganese-enhanced 3D MRI

This magnetic resonance imaging (MRI) study describes mapping of the habenulo-interpeduncular pathway in living mice based on manganese-induced contrast. Six hours after intracerebroventricular microinjection of MnCl2, T1-weighted 3D MRI (2.35 T) at 117 mum isotropic resolution revealed a continuous pattern of anterograde labeling from the habenula via the fasciculus retroflexus to the interpeduncular nucleus. Alternatively, the less invasive systemic administration of MnCl2 allowed for monitoring of the dynamic uptake pattern of respective neural components with even higher reproducibility across animals. Time courses covered the range from 42 min to 24 h after injection. In conclusion, manganese-enhanced MRI may open new ways for functional assessments of the habenulo-interpeduncular system in animal models with cognitive impairment.

Serotonin 5‐HT4 receptors and their mRNAs in rat and guinea pig brain: Distribution and effects of neurotoxic lesions

Serotonin 5-HT4 receptors are widely distributed in the periphery and in brain, where they modulate the release of various neurotransmitters and have been implicated in learning and memory. Nine C-terminal splice variants of this receptor have been cloned in mammalian species. In the rat, three such variants have been described: 5-HT4(a), 5-HT4(b), and 5-HT4(e). In the present study, we have examined several aspects of the distribution of these receptors in brain. First, we provide, in rat and guinea pig, a detailed comparison of the distribution of 5-HT4 receptors labeled by the antagonist [125I]-SB 207710 with the distribution of their encoding mRNA visualized by in situ hybridization histochemistry (ISHH). The results suggest that, in several projection systems (striato-nigral and striato-pallidal pathways, projection from dentate granule cells to field CA3, habenulo-interpeduncular pathway), 5-HT4 receptors are located both somatodendritically and axonally. Second, we have analyzed the distribution of mRNA for the three known rat splice variants by reverse transcription-polymerase chain reaction (RT-PCR) and by ISHH. RT-PCR indicates that all three variants are widely distributed, with 5-HT4(b) mRNA being present in all regions examined (olfactory tubercle, striatum, hippocampus, inferior colliculus, substantia nigra, parietal cortex) and 5-HT4(a) and 5-HT4(e) showing a somewhat more restricted distribution. In other regions (periaqueductal gray, reticular formation, medial septum, diagonal band), faint ISHH signals are observed for 5-HT4(a)+4(e) mRNAs, whereas 5-HT4(b) mRNA signals are almost undetectable. Finally, neurotoxic lesions of basal ganglia components in guinea pig also indicate a location of these receptors on terminals of striatal projection neurons.

Preferential Metabolic Activation of Subcortical Brain Areas by Acute Administration of Nicotine to Rats

Cerebral metabolic and behavioral effects of acutely administered nicotine were measured in rats in relation to dose. Nicotine 0.1, 1, or 10 mg/kg or vehicle was administered intraperitoneally to 3-month-old male Fischer-344 rats that had been pretreated with hexamethonium bromide 5 mg/kg i.p. to reduce peripheral autonomic effects. Regional CMRglc (rCMRglc) values were measured, using the quantitative autoradiographic [14C]-2-deoxy-D-glucose method, in 71 brain regions, beginning 3 min after nicotine or vehicle administration. Intensity of body tremor, scored by a blinded rater, was dose related and peaked at 3 min after nicotine injection. rCMRglc rose in a dose-related manner: Nicotine 0.1 mg/kg had no significant effect in any region, whereas 1 mg/kg elevated rCMRglc significantly in 21 regions (mean rise 20%) and 10 mg/kg produced generalized (56 regions) and greater (mean rise 50%) increases in rCMRglc. Nicotine 1 mg/kg activated thalamic nuclei, cerebellum, geniculate nuclei, superior colliculus, median raphe, reticular formation, and the habenulointerpeduncular pathway, but was without effect in the telencephalon. Effects of nicotine in the hindbrain were related anatomically to reported distributions of [3H]nicotine and [3H]acetylcholine but not [125I]alpha-bungarotoxin binding sites, implying that the former ligands label functional nicotine receptors. The pattern of change in rCMRglc after nicotine administration suggests that its cognitive effects in humans are due to augmented arousal/attention and visual processing rather than to direct neocortical or hippocampal activation.

Cytoarchitecture, fiber connections, and some histochemical aspects of the interpeduncular nucleus in the rat

The organization of afferent and efferent connections of the interpeduncular nucleus (IP) has been examined in correlation with its subnuclear parcellation by using anterograde and retrograde tracing techniques. Based on Nissl, myelin, and acetylcholinesterase staining five paired and three unpaired IP subnuclei are distinguished. The unpaired division includes the rostral subnucleus (IP-R), the apical subnucleus (IP-A), and the central subnucleus (IP-C). The subnuclei represented bilaterally are the paramedian dorsal medial (IP-DM) and intermediate subnuclei (IP-I) and the laterally placed rostral lateral (IP-RL), dorsal lateral (IP-DL), and lateral subnuclei (IP-L). Immunohistochemical techniques showed cell bodies and fibers and terminals immunoreactive for substance P, leu-enkephalin, met-enkephalin, or serotonin to be differentially distributed over the different IP subnuclei. Substance P-positive perikarya were found in IP-R, enkephalin neurons in IP-R, IP-A, and the caudodorsal part of IP-C, and serotonin-containing cell bodies in IP-A and the caudal part of IP-L. Efferent IP projections were studied both by injecting tritiated leucine in IP and by injecting HRP or WGA-HRP in the presumed termination areas. The results indicate that the major outflow of IP is directed caudal-ward to the median and dorsal raphe nuclei and the caudal part of the central gray substance, i.e., the dorsal tegmental region. The projection appears to terminate mainly in the raphe nuclei, around the ventral and dorsal tegmental nuclei of Gudden, and in the dorsolateral tegmental nucleus. The descending projection to the dorsal tegmental region originates in virtually all IP subnuclei, but the main contribution comes from IP-R and the lateral subnuclei IP-RL, IP-DL, and IP-L. Sparser projections to the dorsal tegmental region originate in IP-C and IP-I, whereas the contribution of IP-A is only minimal. The projections from IP-R are mainly ipsilateral and those from IP-DM are mainly contralateral. IP fibers to the median and dorsal raphe nuclei originate predominantly in IP-R and IP-DM, and to a lesser extent in IP-C, IP-I, IP-RL, and IP-DL. A much smaller contingent of IP fibers ascends to diencephalic and telencephalic regions. A relatively minor projection, stemming from IP-RL and IP-DL, reaches the lateral part of the mediodorsal nucleus, the nucleus gelatinosus, and some midline thalamic nuclei. These IP fibers follow either the habenulo-interpeduncular pathway or the mammillothalamic tract.(ABSTRACT TRUNCATED AT 400 WORDS)

The action of cholinomimetic substances on impulse conduction in the habenulointerpeduncular pathway of the rat in vitro.

The effects of some cholinomimetic substances and their antagonists on the peak height of compound action potentials recorded from the terminal region of the habenulointerpeduncular pathway have been studied using a rat brain slice preparation. Carbachol and acetylcholine (ACh) depressed the peak height of the compound action potential and increased the latency to peak. The nicotinic agonists nicotine and dimethylphenylpiperazinium depressed the peak height of the compound action potential while muscarine and glutamate had no effect. The depressant effect of carbachol was blocked by the nicotinic antagonists hexamethonium, mecamylamine and d-tubocurarine but not by atropine. Physostigmine enhanced the effects of ACh and, to a lesser extent, carbachol. In the presence of physostigmine, carbachol or ACh initiated a spontaneous oscillation of the amplitude of the compound action potential which was Ca2+ dependent and was blocked by mecamylamine. It is concluded that depression of the amplitude of the compound action potential is due to activation of presynaptic nicotinic receptors. The results are discussed with reference to possible cholinergic mechanisms in the habenulointerpeduncular pathway.

Acetylcholinesterase Histochemistry of the habenulo-interpeduncular pathway in the rat and the effects of electrolytic and kainic acid lesions.

The histochemical distribution of acetylcholinesterase (AChE) was studied in the habenulo-interpeduncular pathway of normal rats and after electrolytic and kainic acid lesions of the habenular nuclei. From these combined observations it appears that the AChE-rich projection to the interpeduncular nucleus derives from both the medial and the lateral habenular nuclei. The lateral nucleus of the habenula is the main source of AChE-rich fibres in the fasciculus retroflexus, and a number of stained fibres also derive from the stria medullaris. While total habenular lesion completely deprived the fasciculus retroflexus of AChE-stained fibres, a direct effect on the enzyme distribution in the interpeduncular nucleus was only apparent as its rostral pole. In the remainder of the nucleus the AChE distribution did not undergo obvious changes in comparison with the normal pattern, except for a moderate decrease in overall reaction intensity in cases with subtotal habenular lesion bilaterally. The above results are consistent with the observation derived from experiments involving kainic acid injection into the habenula. The neurotoxic effect of kainic acid was highly selective for specific types of neurons in the lateral habenula, while the neurons of the medial habenula were completely unaffected. The existence of an AChE-rich projection from the lateral habenula to the interpeduncular nucleus was supported by a corresponding decrease in enzyme activity in the lateral habenula and fasciculus retroflexus after kainic acid treatment.