Synapses in the interpeduncular nucleus: electron microscopy of normal and habenula lesioned rats.

Four types of synapses have been identified in the rat interpeduncular nucleus. The S synapses were formed by the mainhorizontal plexus of habenulointerpeduncular axons. They contained spherical synaptic vesicles and formed asymmetrical en passant contacts with dendritic processes. The crest synapses had all of these features in common with the S synapses, but occurred in pairs with the two contact zones coextensive and largely parallel on opposite sides of markedly attenuated dendritic processes. Synaptic glomeruli found in this nucleus consisted of several crest synapses engulfed by sheet-like dendritic processes. Identical crest synapses also occurred on dendritic crests. In a limited number of cases, both S and crest synapses arose in common from single axons. Following destruction of the habenular nuclei unilaterally or bilaterally, as expected from the above observations, both S and crest synapses underwent dense degeneration. In some cases with bilateral lesions and in all cases with unilateral lesions, only one of the two axonal endings forming a crest synapse degenerated while the other remained unaffected. Axons containing flat vesicles forming symmetrical axodendritic synapses, axosomatic synapses, and subjunctional bodies associated with S or crest synapses were all minor features of this nucleus. Aspects of the unusual synaptology of this nucleus are discussed in terms of morphology, physiology and transmitter chemistry.

Postnatal synaptogenesis in the rat interpeduncular nucleus.

The rat interpeduncular nucleus (IPN) was studied by electron microscopy from the day of birth to 33 days of age. The habenulointerpeduncular (H-IPN) axons, the principal afferents to IPN, were prominent at birth, occurring in large groups. They formed occasional S synapses en passant at this time. S synapses subsequently increased progressively in number and maturity until 28 days of age. Crest synapses, also formed by the H-IPN axons, appeared at eight days without a recognized formative stage. They exhibited their diverse adult morphology by 14 days of age. F synapses, endings of uncertain origin that contain flattened vesicles, were seen at 21 days, although endings containing small vesicles at 14 days represented the same population. Axosomatic synapses were first seen at four days and remained infrequent subsequently. Dendritic growth cones had their maximal prevalence at birth, decreased at four days and were absent by six days of age. Thus each of the four synaptic types previously described in the adult rat IPN exhibited a characteristic time of appearance and pattern of development in normal rat pups. This well-ordered sequence of synaptogenesis makes the H-IPN system well suited for a study of synaptic plasticity.

Effect of neonatal deafferentation on synaptogenesis in the rat interpeduncular nucleus.

Changes in the normal synaptogenetic pattern of the interpeduncular nucleus (IPN) were induced by destruction of one or both habenular nuclei in neonatal rats. The S synapses, the principal habenulointerpeduncular (H-IPN) afferents, were reduced in number and delayed in time of appearance by unilateral lesions. They were eliminated by bilateral lesions. The crest synapses, also of habenular origin, appeared at ten days of age and were apparently normal in structure after unilateral lesions. However, this actually represented a change in connectivity, since normal crest synapses are of bilateral origin. Following bilateral habenular lesions abnormal crest synapses appeared at 28 days of age. These had normal postsynaptic structure, but presynaptically were formed by non-habenular axons, usually of two different types of each crest synapse. Lesions induced an early appearance of F synapses, at three days, instead of the normal time after six days of age. Apparently similar processes containing flattened vesicles formed somatic contacts only in lesioned animals beginning at 21 days of age. The axosomatic synapses which were only occassional in the adult appeared at the normal time in lesioned animals. At 28 days of age in both unilaterally and bilaterally lesioned animals there was an increase in the number of somatic synapses. This increase was due to an increase of the normal somatic endings plus the appearance of somatic synapses containing flattened vesicles. Both types of somatic contacts were found in similar numbers at subsequent ages up to three months. The changes in S synapses represent Wallerian degeneration. The formation of unilaterally derived crest synapses in unilaterally lesioned animals is interpreted as evidence for a specific recognition mechanism. A postsynaptic control mechanism is proposed to explain the formation of heterologous crest synapses, with the usual distinctive geometry but involving abnormal, non-habenular presynaptic elements four weeks following neonatal bilateral lesions. The increase in axosomatic synapses is unusual since the dendrites of IPN neurons were deafferented, not their somata. The factors leading to the changes in somatic synapses are discussed. The findings reveal new examples of the complexity of neuronal adaptation to CNS lesions in early life.

Modification of left-right pairing during the development of individual crest synapses in the rat interpeduncular nucleus.

The synaptic organization of the rat interpeduncular nucleus is highly ordered in the normal adult. By 90 days of age, 90% of crest synapses in its intermediate subnuclei are formed by two cholinergic endings, one from each medial habenula. Stereological calculation of the number of crest synapses per intermediate subnucleus, based on total samples of crest synapses in 3-4 sections through the subnucleus, allows comparisons of afferent pairing among ages without interference by other developmental changes. Between 21 and 90 days of age, the total number of crest synapses per intermediate subnucleus increases tenfold (p less than 10(-8], from 90,000 at 21 days of age, through 130,000 at 28 days, 440,000 at 45 days, to 1,000,000 at 90 days. The volume of the intermediate subnucleus increases fivefold during the same interval. Electron microscopic degeneration was used to estimate the pairing of left and right habenula afferents at crest synapses at the same ages. Through 21 days of age, only one-third of crest synapses are formed with pairing of one left and one right medial habenula afferent, whereas two-third have both afferent endings arising from the same medial habenula. At 28 days of age left-right pairing has increased to 43%, and at 45 days of age 53%, or 240,000, are so paired. The number of same-side paired crest synapses at 45 days, 210,000, is 3.5 times the number so paired at 21 days (p = .003). This indicates continued formation of considerable numbers of crest synapses with this transient mode of airing as late as 45 days of age.(ABSTRACT TRUNCATED AT 250 WORDS)

The subnuclear organization of the rat interpeduncular nucleus: a light and electron microscopic study.

The subnuclear organization of rat interpeduncular nucleus (IPN) has been examined by light microscopy following staining with Nissl and Holmes methods, 3H-leucine autoradiography, acetylcholinesterase (AChE), and cytochrome oxidase histochemistry on plastic sections stained with toluidine blue, and by electron microscopy. Three unpaired and four paired subnuclei are recognized. The rostral subnucleus is heavily stained for AChE, which clearly delineates its borders. It is distinguished ultrastructurally by two types of synapses on dendrites, and two on perikarya. Of the former, one type is formed by presynaptic processes which contain spherical and dense-cored vesicles and make asymmetrical contacts. Dense-cored vesicles are observed in many of the postsynaptic dendrites. A second type has presynaptic processes containing small, pleomorphic vesicles which make symmetrical contacts. Synapses on perikarya are found in the rostral, central, intermediate, lateral, and interstitial subnuclei. The dorsal subnucleus is continuous with the serotonin-containing B8 cells. The central subnucleus is distinguished by longitudinally oriented medial habenular axons separating palisades of cell bodies. These axons, which also traverse the intermediate subnuclei, form en passant S synapses with small dendrites of the central subnucleus. The intermediate subnuclei react faintly for AChE and intensely for cytochrome oxidase. They contain crest synapses formed by two habenular afferents, one from each medial habenula, which contact a narrow dendritic process en passant. The lateral subnuclei react intensely for AChE and have ultrastructural features similar to the rostral subnuclei. The interstitial subnuclei lie within each fasciculus retroflexus as it enters IPN. The small dorsal lateral subnuclei are evident by light microscopy.

The subnuclear distribution of substance P, cholecystokinin, vasoactive intestinal peptide, somatostatin, leu-enkephalin, dopamine-beta-hydroxylase, and serotonin in the rat interpeduncular nucleus.

The distribution of immunofluorescent somata and processes within the interpeduncular nucleus (IPN) containing substance P (SP), cholecystokinin (CCK), vasoactive intestinal peptide (VIP), somatostatin (SST), leu-enkephalin (L-ENK), dopamine beta hydroxylase (DBH), and serotonin (5HT) was examined in male rats treated with colchicine 48 hours prior to perfusion. Serial sections were examined for immunofluorescence and variations in the density of fluorescence rated 1 + (sparse) to 4 + (dense). The rostral subnucleus contained SP, SST, and L-ENK-positive somata and processes. Substance P and VIP processes were present throughout the rostral subnucleus but were concentrated in two ovoid areas located dorsally in the caudal region of this subnucleus. Cholecystokinin and L-ENK processes surrounded these ovoid areas. The entire width of the central subnucleus was crossed by SP and L-ENK processes oriented horizontally in narrow bands. Substance P processes were also aligned into vertical columns adjacent to the lateral margins of the central subnucleus. Leu-enkephalin and 5HT processes were distributed throughout this subnucleus, while VIP processes were present only caudally. Dopamine beta hydroxylase processes were evenly distributed but were restricted from the vertical columns laterally. The intermediate subnuclei contained a sparse density of SP and 5HT processes that were present in proximity to the major blood vessels penetrating this subnucleus. Only DBH processes were evenly distributed. The lateral subnuclei contained a dense concentration of SP processes. The medial edges of this subnucleus were distinguished by VIP, CCK, L-ENK, and 5HT processes. The dorsal subnucleus contained 5HT, L-ENK, and SST-positive somata and processes. Substance P, VIP, CCK, and DBH processes were also present. Dorsal-lateral subnuclei contained SP, SST, L-ENK, and DBH processes. Interstitial subnuclei contained SP, CCK, L-ENK, 5HT, and DBH processes. This study demonstrates that perikarya and processes containing peptides and monoamines are distributed within subnuclei of IPN in a topographic and heterogeneous pattern. New features of IPN organization are revealed.

Exacerbation generalized nonconvulsive seizures with ethosuximide therapy.

We describe a patient whose absence seizures increased greatly in number after the addition of ethosuximide to ongoing phenobarbital therapy. This unusual response and review of other cases in the literature suggested the possibility of some as yet underfined interaction between ethosuximide and other anticonvulsant drugs. That a particular subytpe of epilepsy may be worsened by ethosuximide therapy is also considered.

Clinical utility of unbound antiepileptic drug blood levels in the management of epilepsy.

We tabulated all unbound and total antiepileptic blood levels collected in 13 months. According to strict criteria, 24% of phenytoin and 15% of valproate unbound blood levels but none of the carbamazepine unbound levels had clinical significance in the management of seizures or side effects. These data support frequent use of unbound phenytoin or valproate blood levels in the management of epilepsy.

Left-right pairing at the crest synapses of rat interpeduncular nucleus.

Synaptic terminals of the habenulo-interpeduncular axons in rat interpeduncular nucleus are demonstrated by electron-microscopic autoradiography following [3H]leucine injection into the habenula. The endings are of two types, S and crest synapses. Previous descriptions of the S synapses based on indirect identification were confirmed. The crest synapses in the interpeduncular nucleus consist of two habenular afferents contacting a narrowed dendrite. Injection of both habenulae with radiolabeled amino acid resulted in autoradiographic labeling of both axons at 48% of individual crest synapses, and of one axon at 39%. However, when only one habenula was injected, no more than one axon was labeled at individual crest synapses; this occurred at 59%. Interpretation of unlabeled endings is possible using the "detection rate" of the method (per cent ratio of labeled to total endings). The detection rate, which can be determined for the bilateral injection case, allows recalculation of the raw data for the unilateral injection case. Left-right pairing of habenular afferents actually occurs at approximately 90% of crest synapses. The use of detection rates in this way, additionally exemplified by analysis of an electron-microscopic degeneration study, is also important in many other applications of both of these experimental neuroanatomical methods, especially when comparisons between two sets of data are made. The conclusions of studies which have not taken this problem into account must be interpreted with caution.

Behavioral and metabolic alterations in the opiate withdrawal syndrome induced by lesions of fasciculus retroflexus.

The interpeduncular nucleus (IPN) appears to be an important integrative center within the limbic system based on its extensive afferent and efferent connections and the presence of numerous neurotransmitters and peptides. Opiate receptors are present within particular subregions of IPN, which is one of the limbic structures showing an increase in regional glucose utilization (RGU) during withdrawal of morphine-addicted rats. The possible role of neural connections in withdrawal was studied by lesioning the main afferent pathway to IPN, the fasciculus retroflexus (FR) bilaterally. Four subnuclei of IPN, lateral, central, rostral and intermediate and FR showed significantly smaller increases in RGU during naloxone-induced withdrawal when compared to sham-operated controls. No difference was found in the apical, dorsal medial or dorsal lateral subnuclei. This metabolic effect of the lesions is not related in any simple way to the localization of opiate receptors or other neurochemical features of IPN. The lesioned animals also had greater weight loss due to diarrhea during withdrawal, consistent with IPN's presumed connection to the vagal nuclei. IPN appears to exhibit local and independent effects of FR lesions during opiate withdrawal.

Neurological assessment of patients on prolonged methadone maintenance.

Medical histories, neurological examinations and electroencephalograms of 25 methadone-maintained subjects were compared with those of 25 abstinent controls. Comparisons on all measures failed to show significant differences between groups. The incidence of abnormalities was low for all subjects. None of the observed conditions appeared to involve methadone in their etiology.

Fornix afferents to the anteroventral thalamic nucleus: an EM study in the rat.

Three types of synapses occur in the anteroventral thalamic nucleus (AVN). Type 1 consists of small (0.5-0.8 microns) axonal endings densely packed with spherical synaptic vesicles. They form markedly asymmetrical synaptic contacts with distal portions of dendrites. Degenerative changes in these axons following destruction of the fornix identify them as the endings of the subicular projection to AVN. Type 2 synapses consist of large (1.0-1.5 microns) axonal processes containing spherical vesicles which form asymmetrical synapses on more proximal dendrites. Type 3 endings consist of large unidentified processes containing spherical, and occasionally flattened, synaptic vesicles forming symmetrical contacts with the largest stem dendrites. Neither of these synaptic types were modified by fornix lesions. The synaptic arrangements within AVN are simpler than other thalamic nuclei in that serial synapses and synaptic glomeruli are not present.

Neurogenesis in subnuclei of the rat interpeduncular nucleus and medial habenula.

Development of the rat habenulo-interpeduncular system is of interest because of its highly ordered adult structure, including seven subnuclei within the interpeduncular nucleus (IPN), localization of synapses of different types and medial habenular (MH) sites of origin within the subnuclei, and differential localization of neurochemicals amongst the subnuclei. In order to further investigate the mechanisms by which these features are produced, the birthdays of neurons within IPN and MH were investigated. All IPN neurons were formed on embryonic days (E) 12 to 16. Birthdays varied for the subnuclei of IPN with the earliest being the serotonin containing apical subnucleus (p less than 0.0001) and the latest being the rostral subnucleus (p less than 0.0001). Rostral lateral and lateral subnuclei were approximately simultaneous with apical, and the intermediate and central subnuclei, related to each other in a number of other ways, had simultaneous birthdays earlier than the rostral subnucleus (p less than 0.0001) and later than the other subnuclei (p less than 0.0001). The medial habenula was found to consist of three regions, termed medial, lateral and dorsal, whose neurons undergo their final mitoses on E15-16, 16-17 and 17-18, respectively. These findings provide additional support for the validity of IPN subnuclei as currently delineated. They suggest mechanisms of development which involve early interactions between specific groups of MH neurons and specific groups of IPN neurons. It is proposed that a sequence of several control mechanisms operates during development to produce this complex system.

Hemicholinium-3 binding sites in subnuclei of the rat interpeduncular nucleus: quantitative in vitro autoradiography.

The interpeduncular nucleus (IPN) receives dense cholinergic input from the medial habenulae (MH) via the fasciculus retroflexus (FR). This projection is known to terminate in the rostral, central and intermediate subnuclei. Correspondingly, the concentration of hemicholinium-3 (HC-3) binding sites in these subnuclei was equal to or greater than that reported in any other brain areas. Moderate values in the distal FR and in the lateral subnuclei indicate that choline uptake sites are located on nonterminal portions of MH afferent axons as well. Possible relationships of HC-3 binding to the unusual metabolic properties of FR and IPN, and to the distribution of choline acetyltransferase-containing axons and terminals in FR and IPN are suggested.

Subdivisions of the interpeduncular nucleus: a proposed nomenclature.

A compromise terminology for subdivisions is proposed which integrates the two previous standard and two recent and more detailed subnuclear schemes. The unpaired median subnuclei are called rostral, central and apical. The paired subnuclei are called rostral lateral, dorsal lateral, intermediate and lateral. A plea is made for attention to these consistent subdivisions, and for uniformity of terminology regardless of the parameters studied.

Quantitative demonstration of somatic synapse sprouting following dendritic deafferentation in neonatal rat interpeduncular nucleus.

Neonatal habenular lesions deafferent the dendrites of interpeduncular nucleus (IPN) neurons by preventing formation of S and crest synapses. The small number of synapses normally contacting IPN neuronal perikarya are of unknown, but non-habenular, origin. The number of synapses on IPN perikarya is significantly increased (p less than 0.001) when 10 control animals are compared to 7 animals with unilateral habenular lesions and 7 animals with bilateral lesions. The increases with bilateral lesions are approximately twice those resulting from unilateral lesions (p less than 0.01). This phenomenon involves sprouting of both the somatic synapses which contain spherical vesicles and those with flattened vesicle endings, and is greater when the latter are considered alone (p less than 0.001). Only the small group of somatic synapses with asymmetrical contacts failed to show a change. The increases suggest a postsynaptic control mechanism which acts in the direction of preserving synaptic input, but permits displacement of the site of input from dendrites to soma. Factors which may be important in determining this outcome are the production of the lesions prior to synaptogenesis, and the presumed shrinkage of the dendrites of the IPN neurons.

In vivo 1H magnetic resonance spectroscopic measurement of brain glycine levels in nonketotic hyperglycinemia.

Nonketotic hyperglycinemia (NKH) is an autosomal recessive disorder of glycine metabolism. Defective glycine cleavage causes elevated concentrations of glycine in plasma, urine, and cerebrospinal fluid. A longitudinal study using magnetic resonance imaging (MRI) and single-voxel 1H magnetic resonance spectroscopy (MRS) was performed on an infant with the typical clinical picture of NKH. He was examined twice during the course of treatment with sodium benzoate and dextromethorphan. At the age of 10 months, MRI showed normal brain structure, while MRS detected a prominent glycine peak in the brain. Repeat MRS at the age of 13 months showed a small increase in glycine peak and a prominent glutamate/glutamine peak not previously detected. The MRS measurements were consistent with the slight increase in blood glycine level and the elevation in glutamine level, indicating that 1HMRS can be a valuable tool in the diagnosis and monitoring of treatment effects in patients with NKH.

Development of the interpeduncular nucleus in the midbrain of Rhesus monkey and human.

The development of the interpeduncular nucleus (IPN) in primates was studied in rhesus monkey with 3H-thymidine autoradiographic, Nissl and Golgi methods and in humans in histological preparations from embryos and fetuses of different ages. Autoradiographic analysis demonstrated that the neurons of the monkey IPN underwent their final cell division between postconception day 36 (E36) and E42, which corresponds to Stages 17 through 21 of Hendrickx and Sawyer. Autoradiograms of monkeys sacrificed at various short intervals following exposure to a pulse of 3H-thymidine showed that IPN neurons were generated in the proximity of the ventricular surface near the confluence of the 3rd ventricle and cerebral aqueduct, migrated ventrally along the midline and then spread laterally after reaching the ventral midbrain, where IPN was first recognized at E45 (Stage 23). The distribution of successively generated neurons in autoradiograms revealed caudal to rostal and lateral to medial spatiotemporal gradients. Differentiation of IPN neuronal size and development of Nissl substance began in rhesus monkey only after postmitotic cells had reached their destination and seemed to be pronounced mainly through E104. However, growth of the dendrites and elaboration of their side branches as seen in Golgi impregnations progressed gradually from E81 to birth (E165) and perhaps even later. Analysis of histological preparations of a series of human embryos and fetuses was used to derive similar information indirectly, since the autoradiographic method cannot be applied to man. It was found that IPN neurons in human probably underwent their final division between Carnegie Stage 17 and 21. Similarly, as in monkey, postmitotic cells in human IPN displayed an inverted fountain pattern of cellular migration. IPN could first be delineated at Stage 23. There was evidence for both caudal to rostral and lateral to medial spatiotemporal gradients in the human, as in the monkey. Thus, in monkey and human, all IPN neurons are generated within the first quarter of intrauterine life and there is remarkable similarity in the timing, tempo and pattern of IPN neuronal differentiation in both species, indicating the validity of using nonhuman primates as an experimental model for understanding the development of this structure in man.

Chronic paroxysmal hemicrania in a young child: possible relation to ipsilateral occipital infarction.

We report a child with chronic paroxysmal hemicrania beginning at 3 years of age with more than 2 years of daily episodes. Indomethacin produced total relief. An ipsilateral, occipital hemorrhagic infarction, probably predating the headaches, may have contributed to their pathogenesis. This is an unusually early onset and persistent chronic paroxysmal hemicrania of possible symptomatic type.

HLA-DR2 in childhood narcolepsy.

A 6-year-old boy with behavioral changes was found to have clinical and electrographically defined narcolepsy. He has HLA-DR2 as do virtually all adult narcoleptics. This finding raises new diagnostic, etiopathogenetic, and future therapeutic issues.