Serotonin-1A autoreceptor binding in the dorsal raphe nucleus of depressed suicides

Serotonergic dysfunction is present in mood disorders and suicide. Brainstem 5-HT1A somatodendritic autoreceptors regulate serotonin neuron firing but studies of autoreceptor binding in the dorsal raphe nucleus (DRN) in depressed suicides report conflicting results. We sought to determine: (1) the anatomical distribution of 5-HT1A receptor binding in the DRN in depressed suicides and psychiatrically normal controls; and (2) whether sex differences in 5-HT1A binding in the DRN contribute to differences between depressed suicides and controls. Previously collected quantitative receptor autoradiograms of [3H]8-hydroxy-2-(di-n-propyl)aminotetralin (3H-8-OH-DPAT) in postmortem tissue sections containing the DRN from drug-free suicide victims (n=10) and matched controls (n=10) were analyzed. Less total receptor binding (fmol/mg tissuexmm3) was observed in the entire DRN in depressed suicides compared with controls (p<0.05). Group differences along the rostrocaudal extent of the DRN were observed for cross-sectional 5-HT(1A) binding (fmol/mg tissue) and receptor binding (fmol/mgxmm3, p<0.05). Cross-sectional 5-HT1A DRN binding in depressed suicides compared with controls was higher rostrally and lower caudally. The differences between depressed suicides and controls were present in males and females, although females had more binding than males. Less autoreceptor binding in the DRN of depressed suicides may represent a homeostatic response to less serotonin release, increasing serotonin neuron firing. More autoreceptor binding in rostral DRN might contribute to deficient serotonin release in ventromedial prefrontal cortex by lower neuronal firing.     

More tryptophan hydroxylase in the brainstem dorsal raphe nucleus in depressed suicides

Deficient serotonin neurotransmission in suicide is indicated by reduced brainstem serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA), fewer 5-HT(1A) autoreceptors and reduced cortical serotonin transporter binding in suicide victims. Tryptophan hydroxylase (TPH) is the rate-limiting enzyme in the synthesis of 5-HT, and alterations in TPH could explain some of these findings. We sought to determine the amount of TPH immunoreactivity (TPH-IR) in the dorsal (DRN) and median (MRN) raphe nuclei in suicides and controls. Brainstems of suicide victims and controls (n = 11 pairs) were collected at autopsy, matched for age, sex and postmortem interval, frozen and sectioned (20 microm). Immunoautoradiography, using an antibody to label TPH, was performed, slides exposed to film and autoradiograms quantified by a computer-based image analysis system. We examined sections every 1000 microm throughout the whole length of the nucleus, performing statistical analysis only on those subjects for whom the raphe was complete (n = 8 pairs). TPH-IR (microCi/g) was higher in suicides than controls (S: 300.8 +/- 70.8 vs. C: 259.6 +/- 40.7, t = 2.57, df = 7, P = 0.04) in the dorsal raphe nucleus (DRN), and not different between suicides and controls (S: 251.3 +/- 44.2 vs. C: 235.9 +/- 27.4, t = 1.49, df = 7, P = 0.18) in the MRN. DRN TPH-IR was higher in male suicide victims (MS) compared to male controls (MC; MS: 318.4 +/- 54.4 vs. MC: 271.9 +/- 22.5, t = 2.66, df = 6, P = 0.03). The analysis of TPH-IR area and density at each DRN rostrocaudal levels showed higher area and density in suicides compared to controls in the rostral DRN and lower area and density in the caudal DRN. TPH-IR, an index of the amount of TPH enzyme, in the DRN is higher in depressed suicides. More TPH may be an upregulatory homeostatic response to impaired serotonin release or less autoreceptor activation. Alternatively, the serotonin impairment in suicide may be due to hypofunctional serotonin-synthesizing enzyme.     

Cigarette Smoking and Tryptophan Hydroxylase 2 mRNA in the Dorsal Raphe Nucleus in Suicides

Cigarette smoking is associated with suicide and mood disorders and stimulates serotonin release. Tryptophan hydroxylase (TPH2) synthesizes serotonin and is over-expressed in suicides. We determined whether smoking is associated with TPH2 mRNA in suicides and controls. TPH2 mRNA was measured postmortem in the dorsal raphe nucleus (DRN) of controls (N = 26, 17 nonsmokers and nine smokers) and suicides (N = 23, 5 nonsmokers and 18 smokers). Psychiatric history was obtained by psychological autopsy. TPH2 mRNA was greater in suicide nonsmokers than suicide smokers, control smokers and control nonsmokers (p = 0.006). There was more TPH2 mRNA throughout the DRN. Smoking interferes with the TPH2 mRNA increase observed in suicide nonsmokers. The absence of altered TPH2 expression in non-suicide smokers suggests no pharmacological effect of smoking.     

Increased tryptophan hydroxylase immunoreactivity in the dorsal raphe nucleus of alcohol-dependent, depressed suicide subjects is restricted to the dorsal subnucleus

Considerable evidence suggests that alcoholics with co-occurring depressive disorder are at greater risk for developing psychosocial problems particularly suicidal behavior. Moreover, dysfunction in serotonin (5-HT) neurotransmission has been implicated in depression, suicide and alcoholism. In the present study, we measured the levels of tryptophan hydroxylase (TPH), the main synthetic enzyme of 5-HT synthesis, in specific nuclei of the dorsal raphe (DR) in depressed suicide victims with alcohol dependence and matched psychiatrically normal controls. TPH immunoreactivity (IR) was quantified in frozen tissue sections containing the DR from 8 suicide victims with a diagnosis of major depression and alcohol dependence, and 8 psychiatrically normal control subjects by using immunoautoradiographic methods. We found that the levels of TPH-IR were significantly increased by 46% in the dorsal subnucleus of the DR in depressed suicide victims with alcohol dependence when compared with controls. In contrast, TPH-IR levels did not significantly differ in the other DR subnuclei between depressed, alcoholic suicide subjects, and controls. Our results indicate that abnormalities in 5-HT biosynthesis in the brain of depressed alcoholic suicide subjects are restricted within distinct regions of the DR.     

Estrogen selectively increases tryptophan hydroxylase-2 mRNA expression in distinct subregions of rat midbrain raphe nucleus: association between gene expression and anxiety behavior in the open field.

BACKGROUND: Ovarian steroids modulate anxiety behavior, perhaps by regulating the serotonergic neurons in the midbrain raphe nucleus. The regulation of the brain-specific isoform of rat tryptophan hydroxylase (TPH2) by ovarian hormones has not yet been investigated. Therefore, we examined the effects of estrogen and progesterone on TPH2 mRNA in the rat dorsal and median raphe nuclei (DRN and MRN, respectively) and whether TPH2 mRNA levels correlated with anxiety behavior. METHODS: Ovariectomized rats were treated for two weeks with placebo, estrogen or estrogen plus progesterone, exposed to the open field test, and TPH2 mRNA was quantified by in situ hybridization histochemistry. RESULTS: Estrogen increased TPH2 mRNA in the mid-ventromedial and caudal subregions of the DRN and the caudal MRN. Combined estrogen and progesterone treatment did not change TPH2 mRNA relative to ovariectomized controls. TPH2 mRNA in caudal DRN was associated with lower anxiety-like behavior, whereas TPH2 mRNA in rostral dorsomedial DRN was associated with increased anxiety-like behavior. CONCLUSIONS: These results suggest that estrogen may increase the capacity for serotonin synthesis in discrete subgroups of raphe neurons, and reinforce previous observations that different subregions of DRN contribute to distinct components of anxiety behavior.     

The neuroscience of suicide

Various postmortem brain studies have provided evidence that reduced serotonin (5-HT) transmission in the ventrolateral prefrontal cortex (vlPFC) is associated with depression-related suicide. Suicide victims have fewer 5-HT transporter sites and a large number of postsynaptic 5-HT1A and 5-HT receptors in the vlPFC, which are implicated in behavioral inhibition and impulsivity. These could be compensatory changes in response to 5-HT hypofunction in depression and suicide. Selective serotonin reuptake inhibitors (SSRIs) are commonly used for the treatment of depression and suicidal ideation. 5-HT innervation of the PFC arises predominantly from 5-HT neurons in the brainstem dorsal raphe nucleus (DRN). In the DRN of suicide cases, 5-HT1A autoreceptors are increased and the levels of 5-HT biosynthetic enzyme, tryptophan hydroxylase (TPH), are reduced. Reduced 5-HT1A feedback inhibition and increased TPH may reflect compensatory changes in response to 5-HT hypofunction in depression-related suicide. Genetic polymorphisms in TPH, 5-HT transporter (5-HTTLPR allele), and 5-HT2A receptor were examined for their association with depression-related suicide, but no consistent associations were found. Stress is a risk factor for depression and is linked to hyperactivity of the hypothalamo-pituitary-adrenal axis and suicide. Corticotropin-releasing factor (CRF)-immunoreactive varicose fibers were detected in the DRN of suicide victims, suggesting that CRF neurons in the paraventricular nucleus of the hypothalamus and 5-HT neurons in the DRN may form a circuit in stress-induced depression. Alcoholics are at a significantly greater risk of suicide than the general population. Alcoholism is associated with alterations in the 5-HT system.     

Cytoarchitecture of the human dorsal raphe nucleus

Serial 50 microns Nissl-stained sections through the midbrain and pontine central gray of four adult humans (mean age 56 years, mean postmortem delay 3 hours) were analysed and the subnuclei of the dorsal raphe nucleus (DR) delineated on the basis of neuronal morphology and density. Five subnuclei were apparent: the interfascicular, ventral, ventrolateral, dorsal, and caudal. The area of each subnucleus was measured in sections selected at regular intervals throughout the length of the DR. The number of neurons was counted and their density within each subnucleus calculated. The dorsal subnucleus was the largest and contained the majority of neurons but had the lowest neuronal density. The ventrolateral subnucleus had the highest density of neurons. A total of 235,000 +/- 15,000 neurons (average of 1,200 +/- 200 neurons per section) were found within a volume of 71.3 +/- 4.5 mm3 of DR with a mean neuronal density of 3,300 +/- 200 neurons/mm3. Morphometric and morphological analysis of DR neurons revealed four distinct neuron types: round, ovoid, fusiform, and triangular. These types of neurons characterized particular subnuclei. The location and boundaries of the subnuclei of the human dorsal raphe are presented in the form of an atlas. The subdivisions described are similar to that described in other mammals. On the basis of this information the location of particular projection neurons within the human dorsal raphe can be predicted and the effects of disease on this nucleus may be forecast.     

Morphometry of the dorsal raphe nucleus serotonergic neurons in suicide victims.

BACKGROUND: The serotonin deficiency hypothesis of suicide has been important heuristically. Few studies have directly examined the brainstem dorsal raphe nucleus (DRN) serotonin neurons. We determined the number and morphometry of DRN serotonergic neurons in suicide victims (n = 7) compared to controls (n = 6). METHODS: Brainstems were collected at autopsy, fixed and cryoprotected. Tissue was sectioned, stained for Nissl and processed with an antiserum that cross-reacts with tryptophan hydroxylase. All DRN neurons were identified, counted and analyzed every 1000 microns. Neuron morphometry was characterized by soma area (micron 2), sphericity, perimeter, length and density (neurons per mm3). RESULTS: Neuron number and density was higher in suicide victims (1,780 +/- 127 neurons/mm3) than controls (1,349 +/- 68). The DRN volume did not differ between groups (66 +/- 9 mm3 for controls vs. 67 +/- 5 mm3 for suicides). Mean neuronal area and sphericity did not differ between suicides and controls. The total number and the density of DRN neurons did not correlate with age. CONCLUSIONS: The finding of an increased number of neurons indicates that impaired serotonergic transmission found in association with serious suicide attempts is not due to fewer neurons.     

The serotonin transporter in the midbrain of suicide victims with major depression.

BACKGROUND: The involvement of serotonin in depression and suicide has been proposed, because major depression is successfully treated by medications that specifically block the serotonin transporter, and there is evidence for a decrease in serotonin transporters in major depression and suicide. The midbrain dorsal raphe nucleus (DR) has been implicated as a site for diminished serotonergic activity in that suicide victims with major depression have a significant increase in serotonin-1A autoreceptors in the DR. METHODS: [(3)H]Paroxetine was used to label the serotonin transporter in the subnuclei of the DR at several rostral-to-caudal levels of the midbrain in ten pairs of suicide victims with major depression and age-matched psychiatrically normal control subjects. RESULTS: There was a significant increase in serotonin transporters in the entire DR progressing from rostral-to-caudal levels in both normal control subjects and suicide victims with major depression. At comparable rostral-to-caudal levels, there were no significant differences in [(3)H]paroxetine binding between depressed suicide victims and normal control subjects in either the entire DR or its constituent subnuclei. CONCLUSIONS: The pathophysiology of serotonin mechanisms in suicide victims with major depression does not appear to involve alterations in the binding of [(3)H]paroxetine to the serotonin transporter in the midbrain DR.     

Increased mRNA expression of cytochrome oxidase in dorsal raphe nucleus of depressive suicide victims

Suicidal behavior is a problem with important social repercussions. Some groups of the population show a higher risk of suicide; for example, depression, alcoholism, psychosis or drug abuse frequently precedes suicidal behavior. However, the relationship between metabolic alterations in the brain and premorbid clinical symptoms of suicide remains uncertain. The serotonergic and noradrenergic systems have frequently been, implicated in suicidal behavior and the amount of serotonin in the brain and CSF of suicide victims has been found to be low compared with normal subjects. However, there are contradictory results regarding the role of noradrenergic neurons in the mediation of suicide attempts, possibly reflecting the heterogeneity of conditions that lead to a common outcome. In the present work we focus on the subgroup of suicide victims that share a common diagnosis of major depression. Based on post-mortem studies analyzing mRNA expression by in situ hybridization, serotonergic neurons from the dorsal raphe nucleus (DRN) from depressive suicide victims are seen to over-express cytochrome oxidase mRNA. However, no corresponding changes were found in the expression of tyrosine hydroxylase (TH) mRNA in the noradrenergic neurons of the Locus Coeruleus (LC). These results suggest that, despite of the low levels of serotonin described in suicide victims, the activity of DRN neurons could increase in the suicidally depressed, probably due to the over activation of serotonin re-uptake. No alteration was found in noradrenergic neurons, suggesting that they play no crucial role in the suicidal behavior of depressive patients.     

Quantitative mapping of tryptophan hydroxylase-2, 5-HT1A, 5-HT1B, and serotonin transporter expression across the anteroposterior axis of the rat dorsal and median raphe nuclei

Depression and anxiety disorders are among the leading causes of morbidity, mortality, and disability in the United States. Impaired serotonin neurotransmission appears to be a central mechanism inducing depressive and anxiety symptoms. Most serotonergic innervation of the forebrain arises from the median raphe nucleus (MRN) and dorsal raphe nucleus (DRN). The DRN displays a complex internal morphology, with distinct subregions varying across the anteroposterior (A-P) axis. However, many studies have considered the DRN as a whole or used easily confused terminology to describe position. Given the large differences in receptor expression, electrophysiological properties, and connectivity between various subregions of the DRN, it appears probable that they have distinct functional roles in the regulation of behavior. To foster uniform definitions of locations within these nuclei, we have quantitatively mapped gene expression in DRN and MRN for tryptophan hydroxylase-2 (Tph2), the serotonin transporter, as well as 5-HT1A and 5-HT1B receptors. These quantitative studies revealed differences in the density of expression of each gene in the ventromedial, dorsomedial, and dorsolateral subnuclei of the DRN, as well as distinct variation in expression across the A-P axis. These findings provide additional evidence that subregions of the DRN are heterogeneous and need to be considered independently. In addition, a fine scale map of Tph2 expression suggests definitions for categorical divisions of the DRN across the A-P axis. These are based on distinct morphological patterns of Tph2 expression and may be more reflective of physiology than the classic terminology dividing the DRN into equal thirds.     

The Expression of Brain-Derived Neurotrophic Factor and Tryptophan Hydroxylase in the Dorsal Raphe Nucleus during Repeated Stress

The mechanisms of development of stress-induced depression symptoms may include interaction between the elements of the serotonergic (5-HT) system and brain-derived neurotrophic factor (BDNF) at the level of the cell bodies of 5-HT neurons. In order to evaluate this situation, we used immunofluorescent staining of BDNF protein and the enzyme of serotonin synthesis tryptophan hydroxylase (TPH) in the dorsal raphe nucleus (DRN) of adult male rats that were subjected daily to a 15-min forced swim stress for 1, 2, 7, or 14 days. The expression of the BDNF protein was substantially decreased in the DRN after 7- or 14-day stress, whereas the expression of the TPH protein significantly increased after the 14-day stress and did not differ from the control after the shorter duration of repeated stress. The absence of correlation between some indices indicates their independence from each other at the level of the cell bodies of 5-HT neurons.     

Peptidergic and aminergic innervation of the facial nucleus of the rat with special reference to ontogenetic development

The distribution and ontogenetic development of several neuropeptides such as enkephalin, substance P, somatostatin, neuropeptide Y, and of monoamines such as serotonin and catecholamines in the facial nucleus of the rat were investigated with immunocytochemistry. The neuropeptides were concentrated in certain subnuclei. Enkephalin-immunoreactive fibers were distributed in the medial and dorsal subnuclei, substance P in the intermediate and dorsal subnuclei, somatostatin in the intermediate subnucleus, and neuropeptide Y in the dorsal subnucleus. The amines were distributed evenly throughout the nucleus. These distribution patterns suggest that peptidergic fibers are closely related to the functions of different subnuclei, while fibers containing monoamines are more basic--not specific to individual muscles. Few of these fibers were observed in the prenatal stage of the rat, but they increased markedly in number during the first postnatal week, and had established their innervation pattern by the tenth postnatal day, which coincides with the establishment of nerve-muscle innervation. The present study further showed that fibers containing serotonin are supplied mainly from the raphe nucleus, that catecholamine fibers are from neurons containing catecholamine surrounding the facial nucleus, and that fibers containing neuropeptide Y are from the lateral part of the caudal medullary reticular formation. These findings suggest that catecholamine and neuropeptide Y are not both present in the single neurons projecting to the facial nucleus.     

Topographic organization of serotonergic dorsal raphe neurons projecting to the superior colliculus in the Mongolian gerbil (Meriones unguiculatus).

Recent evidence suggests that the dorsal raphe nucleus (DRN) of the brainstem is a collection of neuronal clusters having different neurochemical characteristics and efferent projection patterns. To gain further insight into the neuroanatomic organization of the DRN, neuronal populations projecting to the superior colliculus (SC) were mapped in a highly visual rodent, the Mongolian gerbil (Meriones unguiculatus). Retrograde tracers Fluoro-Gold (FG) or cholera toxin subunit-B (CTB) were injected into the superficial layers of the SC, and serotonin (5-hydroxytryptamine, 5-HT) -positive cells were identified by using immunocytochemistry in the FG-injected animals. Based on its projections to the SC, the DRN was divided into five rostrocaudal levels. In the rostral and middle levels of the DRN, virtually all FG-filled cells occurred in the lateral DRN, and 36-55% of 5-HT-immunoreactive (5-HT-ir) cells were also double-labeled with FG. Caudally, FG-filled cells occurred in the lateral, ventromedial, and interfascicular DRN; and 44, 12, and 31% of 5-HT-ir cells, respectively, were also FG-filled. The dorsomedial DRN contained only a small proportion of FG-filled cells at its most caudal level and was completely devoid of FG-filled cells more rostrally. The CTB-injected animals showed a similar distribution of retrogradely labeled cells in the DRN. Topographically, the dorsal tegmental nucleus and the laterodorsal tegmental nucleus appeared to be closely associated with 5-HT-ir cells in the caudal DRN. These results suggest that the lateral DRN and the ventromedial/interfascicular DRN may be anatomically, morphologically, and neurochemically unique subdivisions of the gerbil DRN.     

Afferent projections to the interpeduncular nucleus in the rat, as studied by retrograde and anterograde transport of wheat germ agglutinin conjugated to horseradish peroxidase

We examined the afferent projections to the subnuclei of the interpeduncular nucleus (IPN) in the rat by means of retrograde and anterograde transport of wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP). We observed locations of retrogradely labeled cells following injections of WGA-HRP into the IPN, and distributions of anterogradely labeled fibers and terminals within the IPN following injections into the areas that contain cells of origin of afferents. Results of the retrograde and anterograde experiments have clarified the detailed organization of the IPN afferents. A part of the nucleus incertus, located dorsomedial to the dorsal tegmental nucleus, projects to the contralateral half of the rostral subnucleus of the IPN; the pars caudalis of the dorsal tegmental nucleus projects sparsely to the rostral lateral, dorsal lateral, lateral, caudal, and apical subnuclei predominantly contralaterally; the laterodorsal tegmental nucleus, to most of the subnuclei predominantly contralaterally; the ventromedial central gray rostral to the dorsal tegmental nucleus and lateral to the dorsal raphe nucleus projects to the rostral lateral and dorsal lateral subnuclei predominantly contralaterally; the median raphe nucleus, substantially to all subnuclei; the medial habenular nucleus, in a topographic manner, to the rostral, central, and intermediate subnuclei, to the rostral lateral and lateral subnuclei predominantly ipsilaterally, and to the dorsal lateral subnucleus predominantly contralaterally; the supramammillary nucleus and areas around the origin of the mammillothalamic tract and near the third ventricle project sparsely to the ventral part of the rostral subnucleus and to the central, lateral, caudal and apical subnuclei; the nucleus of the diagonal band, sparsely to the rostral, central, dorsal lateral, caudal, and apical subnuclei. These differential projections of the afferents to the subnuclei of the IPN may reflect its complex functions within the limbic midbrain circuit.     

鱼藤酮处理大鼠脑内VMAT2及TPH、5-HT的表达改变

目的观察鱼藤酮处理大鼠脑内单胺囊泡转运体2(VMAT2)。色氨酸羟化酶(TPH)及5—羟色胺(5—HT)的表达变化;探讨其对5—HT能神经元的影响及可能机制。方法选用健康、成年雄性Wistar大鼠,背部皮下注射鱼藤酮3d或28d制作大鼠动物模型;利用免疫细胞化学分析VMAT2在大鼠黑质、中缝背核和尾壳核以及TPH、5—HT在中缝背核的表达变化;以透射电镜观察轴突超微结构的改变,探讨鱼藤酮毒性作用的可能机制。结果鱼藤酮3d组:(1)鱼藤酮组大鼠黑质、中缝背核VMAT2的表达增加,免疫反应强度吸光度值显著高于对照组;尾壳核VMAT2的表达减弱,免疫反应强度吸光度值明显低于对照组(P0.01);(2)鱼藤酮组大鼠中缝背核TPH、5—HT的表达减少,免疫反应强度吸光度值显著低于对照组(P0.01);(3)透射电镜观察显示鱼藤酮处理大鼠中缝背核轴突变性、其内微管解聚。鱼藤酮28d组:(1)鱼藤酮组大鼠黑质、中缝背核及尾壳核VMAT2的表达均减弱,免疫反应强度吸光度值显著低于对照组(P0.01或P0.05);(2)与对照组相比,鱼藤酮组大鼠中缝背核TPH、5—HT的表达降低,免疫反应强度吸光度值显著低于对照组(P0.01)。结论鱼藤酮降低5—HT能神经元TPH、5—HT的表达,其毒性作用可能与轴突微管解聚导致突触囊泡VMAT2轴浆顺行转运障碍有关。     

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)