Ultrasonographic and hemodynamic characteristics of patients with symptomatic carotid near-occlusion: results from a multicenter registry study.

PURPOSE: The ultrasonographic and hemodynamic features of patients with carotid near-occlusion (CNO) are still not well known. Our aim was to describe the ultrasonographic and hemodynamic characteristics of a cohort of patients with CNO. METHODS: A prospective, observational, nationwide, and multicenter study was conducted from January/2010 to May/2016. Patients with digital subtraction angiography (DSA)-confirmed CNO were included. We collected information on clinical and demographic characteristics, carotid and transcranial ultrasonography and DSA findings, presence of full-collapse, collateral circulation, and cerebrovascular reactivity (CVR). RESULTS: One hundred thirty-five patients were analyzed. Ultrasonographic and DSA diagnosis of CNO were concordant in only 44%. This disagreement was related to the presence/absence of full-collapse: 45% of patients with CNO with full-collapse were classified as a complete carotid occlusion, and 40% with a CNO without full-collapse were interpreted as severe stenosis (p < 0.001). Mean velocities (mV) and pulsatility indexes (PIs) were significantly lower in the ipsilateral middle cerebral artery compared with the contralateral (43 cm/s vs 58 cm/s, p < 0.001; 0.80 vs 1.00, p < 0.001). Collateral circulation was identified in 92% of patients, with the anterior communicating artery (73%) being the most frequent. CVR was decreased or exhausted in 66% of cases and was more frequent in patients with a poor or absent collateral network compared with patients with ≥ 2 collateral arteries (82% vs 56%, p = 0.051). CONCLUSION: The accuracy of carotid ultrasonography in the diagnosis of CNO seems to be limited, with significant discrepancies with DSA. Decreased ipsilateral mV, PI, and CVR suggest a hemodynamic compromise in patients with CNO.

The risk of recurrent stroke at 24 months in patients with symptomatic carotid near-occlusion: results from CAOS, a multicentre registry study.

BACKGROUND AND PURPOSE: The risk of recurrent stroke amongst patients with symptomatic carotid near-occlusion (SCNO) has not been clearly established, and its management remains controversial. The aim was to define the 24-month risk of recurrent stroke and to analyse the effect of the different treatment modalities (medical treatment and revascularization) in a population of patients with SCNO. METHODS: A multicentre, nationwide, prospective study from January 2010 to May 2016 was performed. Patients with angiography-confirmed SCNO were included. The primary end-point was ipsilateral ischaemic stroke including periprocedural events within 24 months following the presenting event. Revascularization results and periprocedural complications, ipsilateral transient ischaemic attack, disabling or fatal stroke, and mortality were also noted. RESULTS: The study population comprised 141 patients from 17 Spanish centres. Seventy patients (49.6%) were treated by revascularization (carotid stenting in 47, endarterectomy in 23). Complete revascularization was achieved in 58 patients (83%). Periprocedural stroke or death occurred in 5.7%. The 24-month cumulative incidence of the primary end-point was 11.1% (95% confidence interval 5.8-16.4; n = 15), 12% in the medical treatment group and 10.2% in the revascularization group, log-rank P = 0.817. The cumulative rates of ipsilateral ischaemic stroke or transient ischaemic attack, disabling or fatal stroke, and mortality, were 17%, 4.5% and 7.5%, respectively. CONCLUSIONS: The rate of ipsilateral ischaemic stroke in patients with SCNO seems to be lower than the known rate associated with severe carotid stenosis without near-occlusion. The potential benefit of revascularization in the prevention of stroke in patients with SCNO may be influenced by the effectiveness and safety of the procedure.

Visual snow syndrome and its relationship with migraine.

INTRODUCTION: Visual snow syndrome (VSS) is a central nervous system disorder that consists of the constant perception of small black and white dots throughout the entire visual field. DEVELOPMENT: VSS can present from infancy to old age, with greater prevalence in the young population, and shows no difference between sexes. The diagnostic criteria include the presence of visual snow and such other visual phenomena as palinopsia, photophobia, nyctalopia, and other persistent visual phenomena. The pathophysiology of VSS is unknown, but hyperexcitability of the visual cortex and a dysfunction in higher-order visual processing are postulated as potential mechanisms. The prevalence of migraine among patients with VSS is high, compared to the general population, and symptoms are more severe in patients presenting both conditions. No effective treatment is available, but the drug with the best results is lamotrigine, which is recommended only in selected cases with severe functional limitation. CONCLUSIONS: VSS is a little-known and underdiagnosed entity, but the increasing number of studies in recent years has made it possible to establish diagnostic criteria and begin studying its pathophysiology. This entity is closely related to migraine, with overlapping symptoms and probably shared pathophysiological mechanisms.

Endarterectomy, Stenting, or Medical Treatment for Symptomatic Carotid Near-Occlusion: Results from CAOS, a Multicenter Registry Study.

BACKGROUND AND PURPOSE: The treatment of symptomatic carotid near-occlusion is controversial. Our aim was to analyze the results of carotid endarterectomy and carotid artery stent placement in patients with symptomatic carotid near-occlusion and to identify factors related to technical failure, periprocedural complications, and restenosis. MATERIALS AND METHODS: We conducted a multicenter, prospective nonrandomized study. Patients with angiography-confirmed carotid near-occlusion were included. We assessed the revascularization rate and periprocedural stroke or death. Twenty-four-month clinical and carotid imaging follow-up was performed, and rates of carotid restenosis or occlusion, ipsilateral stroke, and mortality were analyzed. Carotid artery stent placement, carotid endarterectomy, and medical treatment were compared. RESULTS: One hundred forty-one patients were included. Forty-four carotid artery stent placement and 23 carotid endarterectomy procedures were performed within 6 months after the event. Complete revascularization was achieved in 83.6%, 81.8% in the carotid artery stent placement group and 87% with carotid endarterectomy (P = .360). Periprocedural stroke or death occurred in 6% (carotid artery stent placement = 2.3%; carotid endarterectomy = 13%; P = .077) and was not related to revascularization failure. The carotid restenosis or occlusion rate was 8.3% (5% restenosis, 3.3% occlusion); with carotid artery stent placement it was 10.5%; and with carotid endarterectomy it was 4.5% (P = .419). The 24-month cumulative rate of ipsilateral stroke was 4.8% in the carotid artery stent placement group, 17.4% for carotid endarterectomy, and 13.1% for medical treatment (P = .223). Mortality was 12%, 4.5%, and 5.6%, respectively (P = .422). Revascularization failure and restenosis occurred more frequently in patients with full collapse compared with patients without full collapse (33.3% versus 5.6%, P = .009; 21.4% versus 2.9%, P = .032, respectively). CONCLUSIONS: Carotid artery stent placement and carotid endarterectomy are associated with high rates of failure and periprocedural stroke. Carotid near-occlusion with full collapse appears to be associated with an increased risk of technical failure and restenosis. Carotid near-occlusion revascularization does not seem to reduce the risk of stroke at follow-up compared with medical treatment.

Visual snow syndrome and its relationship with migraine. / Síndrome de nieve visual y su relación con la migraña.

INTRODUCTION: Visual snow syndrome (VSS) is a central nervous system disorder that consists of the constant perception of small black and white dots throughout the entire visual field. DEVELOPMENT: VSS can present from infancy to old age, with greater prevalence in the young population, and shows no difference between sexes. The diagnostic criteria include the presence of visual snow and such other visual phenomena as palinopsia, photophobia, nyctalopia, and other persistent visual phenomena. The pathophysiology of VSS is unknown, but hyperexcitability of the visual cortex and a dysfunction in higher-order visual processing are postulated as potential mechanisms. The prevalence of migraine among patients with VSS is high, compared to the general population, and symptoms are more severe in patients presenting both conditions. No effective treatment is available, but the drug with the best results is lamotrigine, which is recommended only in selected cases with severe functional limitation. CONCLUSIONS: VSS is a little-known and underdiagnosed entity, but the increasing number of studies in recent years has made it possible to establish diagnostic criteria and begin studying its pathophysiology. This entity is closely related to migraine, with overlapping symptoms and probably shared pathophysiological mechanisms.

[Development of serotonergic neurons of dorsal raphe nuclei in mice with knockout of monoamine oxidase A and 5-HT1A and 5-HT1B autoreceptor].

The morphological changes in the development of serotonergic neurons of the dorsal raphe nuclei in the medulla oblongata was studied by immunocytochemistry in mice with knockout of 1A and 1B serotonin autoreceptors as well as monoamine oxidase A. Serotonin autoreceptors regulate electric activity of serotonergic neurons as well as the synthesis and release of the neurotransmitter, while monoamine oxidase A catalyzes its degradation. These genetic modifications proved to have no effect on the number of serotonergic neurons in the medulla oblongata but induced morphofunctional changes. Decreased cell size and increased intracellular serotonin level were observed in the case of monoamine oxidase A deficiency, while excessive cell size and decreased intracellular serotonin level were observed in the case of autoreceptor deficiency. The data obtained confirm the hypothesis of autoregulation of serotonergic neurons in development.

[Pseudoaneurysm of the hepatic artery: rare complication of chronic pancreatitis]. / Pseudoanevrisme de l'artere hepatique: complication rare au cours de l'histoire naturelle de la pancreatite chronique: à propos de deux cas.

Bleeding from a pseudoaneurysm is a rare complication of chronic pancreatitis. We present two cases of ruptured pseudoaneurysms of the hepatic arteries. The first case involved a pancreatic pseudocyst that ruptured in the duodenum and the second resulted in an intrahepatic hematoma that compressed the bile tract causing secondary hemobilia. Angiographic embolization was the primary treatment in both cases, with surgery for the first patient and later radiological drainage in the second. The hemorrhage was controlled in both cases despite the severe prognosis and high mortality in these cases.

Acute fatal pulmonary embolism during cyanoacrylate injection in gastric varices.

We report a case of massive pulmonary embolism during cyanoacrylate glue endoscopic injection in a patient with gastric varices from portal hypertension. A review of the literature and results in an animal model show the physiopathology and risk factors associated with this endoscopic procedure.

[Fibrinolytic treatment with tissue plasminogen activator. Outcomes in clinical practice with a multidisciplinary model of intervention]. / Tratamiento fibrinolítico con activador del plasminógeno tisular. Resultados en la práctica clínica con un modelo de actuación multidisciplinario.

INTRODUCTION AND AIMS: Until now treatment of acute cerebral infarction with tissue plasminogen activator (rt-PA) has been applied to a limited extent in large medical centres with acute stroke units. The aim of this study is to determine the safety and effectiveness of this treatment in a centre with no previous experience following a multidisciplinary model. PATIENTS AND METHODS: We conducted a prospective, observation-based study involving patients who were treated with intravenous rt-PA within 3 hours of suffering an acute ischaemic stroke between January 2004 and December 2006. Basal variables, haemorrhagic complications, response to treatment and functional progress were evaluated using the modified Rankin Scale (mRS) at 3 months. Treatment and control were applied following a multidisciplinary protocol implemented by the Intensive Medicine and Neurology services. RESULTS: In all, 46 patients were treated, their mean age being 67 +/- 12 years (63% males). NIH pre-treatment: 13.6 +/- 4.7; median: 13.5; range: 5-22. Time elapsed between the onset of symptoms and arrival at the hospital was 53 +/- 27 min, and door-to-needle time was 69 +/- 27 min. At 24 hours, 48% of patients had improved their scores on the NIH scale > 4 points. A total of 10 haemorrhagic transformations (21.7%) were observed, none of which were symptomatic. At three months, 54.3% of patients were functionally independent (mRS: 0-2). Mortality rate at 90 days was 13.1%. CONCLUSIONS: Administration of rt-PA to patients with acute ischaemic stroke in health care practice following a multidisciplinary protocol is safe and has a neurological progression that is comparable to clinical trials and studies.

Identification of lumbar spinal neurons controlling simultaneously the prostate and the bulbospongiosus muscles in the rat.

Lumbar spinothalamic neurons in the lamina X of the L3-L4 spinal cord segment have been proposed to constitute the spinal ejaculation generator in male rats. Lumbar spinothalamic cells are immunoreactive for galanin and neurokinin-1 receptors. We previously showed that after injection of pseudorabies virus either in the bulbospongiosus muscle or in the prostate, retrogradely labeled cells in the L3-L4 segment also displayed galanin or neurokinin-1 receptor immunoreactivities, demonstrating a direct link between lumbar spinothalamic cells and two anatomical structures involved in the two phases of ejaculation i.e. the emission and the expulsion phases. In order to provide with a more precise anatomical support for the role of lumbar spinothalamic cells in controlling ejaculation, we injected simultaneously in male adult rats two strains of recombinant pseudorabies virus, expressing either beta-galactosidase (PRV-BaBlu) or green fluorescent protein (PRV-152) in the prostate and in the bulbospongiosus muscle, respectively. After 5 days, we performed multiple immunofluorescence experiments to detect PRV-BaBlu, PRV-152 and galanin or neurokinin-1 receptors in transverse sections of the L1-S1 segment. Double- and triple-labeled cells were counted using confocal laser scanning microscope. Double-labeled neurons with the two strains of pseudorabies virus were mainly found at the L3-L4 segment lateral to the central canal in lamina X and represented about 60% of the total number of pseudorabies virus-labeled neurons. All the double pseudorabies virus-labeled neurons also expressed lumbar spinothalamic and most of them neurokinin-1 receptor, identifying them as lumbar spinothalamic neurons. The convergence of retrograde labeling from prostate and bulbospongiosus muscle on the same lumbar spinothalamic cells strongly reinforce their role in the spinal control and coordination of the emission and expulsion of sperm.

Galanin and neurokinin-1 receptor immunoreactive [corrected] spinal neurons controlling the prostate and the bulbospongiosus muscle identified by transsynaptic labeling in the rat.

Ejaculation requires the coordination of sympathetic, parasympathetic and somatic neural outputs. Timely occurrence of the emission and expulsion of sperm results from an interplay between spinal nuclei innervating the seminal tract and the sexual accessory glands including the prostate on the one hand, and on the other hand perineal striated muscles, particularly the bulbospongiosus muscle. A group of cells essential for ejaculation, located around the central canal and referred to as lumbar spinothalamic neurons have been recently identified. Lumbar spinothalamic neurons are immunoreactive for galanin and neurokinin-1 receptor. In order to investigate the anatomical relationships between lumbar spinothalamic neurons and both the prostate and the bulbospongiosus muscle, pseudorabies virus retrograde tracing technique was used combined with immunohistochemistry. Three to five days after pseudorabies virus injection in the bulbospongiosus muscle or the prostate in male rats, spinal cord sections were processed for double immunofluorescence against pseudorabies virus and galanin or neurokinin-1 receptor. Immunocytochemical experiments against pseudorabies virus and choline acetyltransferase were also performed to discriminate between motoneurons and preganglionic neurons, or interneurons. Spinal sections were examined with confocal laser scanning microscope. Three days after pseudorabies virus injection within the prostate and the bulbospongiosus muscle, sympathetic preganglionic neurons and motoneurons of the dorsomedial nucleus were retrogradely labeled, respectively. Five days after pseudorabies virus injection, transsynaptically labeled choline acetyltransferase-negative neurons were found mainly located in the medial gray surrounding the central canal from L1 to S1. At the L3-L4 level, most of transsynaptically labeled neurons were immunoreactive for galanin and to a lesser extent for neurokinin-1 receptor, strongly suggesting that they could be the lumbar spinothalamic cells. We have thus evidenced connections between these cells and motoneurons of the dorsomedial nucleus and both sympathetic and parasympathetic preganglionic neurons innervating the bulbospongiosus muscle and the prostate, respectively. These anatomical data reinforce the crucial role for lumbar spinothalamic cells in coordinating the spinal control of ejaculation.

Aging associated changes in serotoninergic and dopaminergic pre- and postsynaptic neurochemical markers in the rat brain.

Measurements of endogenous levels of serotonin (5-HT), 5-hydroxyindole acetic acid (5-HIAA), dopamine (DA) and dihydroxyphenyl acetic acid (DOPAC), and biochemical and autoradiographic investigations on 5-HT and DA receptors were made in various brain regions in male rats at three different ages: 3 months, 10 months and 22 months. Age-dependent decreases in 5-HT levels associated with parallel increases in 5-HIAA/5-HT ratio were observed in the hypothalamus, striatum, hippocampus and cerebral cortex, suggesting an accelerated 5-HT turnover in aged rats. Similarly, DA levels were lower, and DOPAC/DA ratio was higher in the striatum of 22-month-old compared to 3-month-old or 10-month-old rats. Of the three different classes of 5-HT receptors which were examined, 5-HT1B sites exhibited the largest age-dependent decrease in density, followed by 5-HT2 sites, while 5-HT1A sites remained practically unchanged during aging. By comparison, the loss of striatal D2 receptors in 22-month-old rats compared to young adults was much greater than that of any 5-HT receptor subtype. Such differential age-dependent alterations of the various classes of 5-HT receptors and of dopaminergic versus serotoninergic synaptic markers might be responsible for at least some of the functional deficits in aged animals.

Immunocytochemical localization of serotonin1A receptors in the rat central nervous system.

Specific anti-rat 5-hydroxytryptamine1A (serotonin1A) receptor antibodies raised in a rabbit injected with a synthetic peptide corresponding to a highly selective portion of the third intracellular loop of the receptor protein (El Mestikawy et al. [1990] Neurosci. Lett. 118:189-192) were used for immunohistochemical mapping of serotonin1A receptors in the brain and spinal cord of adult rats. The highest density of immunostaining was found in limbic areas (lateral septum, CA1 area of Ammon's horn and dentate gyrus in the hippocampus, and frontal and entorhinal cortices), in the anterior raphe nuclei, and in the interpeduncular nucleus, in agreement with previous autoradiographic studies with selective radioligands showing the enrichment of these regions in serotonin1A receptor binding sites. Serotonin1A receptor-like immunoreactivity was also present, but at a moderate level, in the neocortex, in some thalamic and hypothalamic nuclei, in the nucleus of the solitary tract, in the dorsal tegmentum, in the nucleus of the spinal tract of the trigeminal nerve, and in the superficial layers of the dorsal horn in the spinal cord. In contrast, extrapyramidal areas, including the caudate putamen, the globus pallidus, and the substantia nigra as well as the cerebellum, exhibited very low to no immunostaining by antiserotonin1A receptor antibodies. At the cellular level, both the plasma membrane of neuronal perikarya and fine neuronal processes probably corresponding to dendritic fields were found to bind antiserotonin1A receptor antibodies. Regional differences were noted regarding these two types of immunostaining, because only dendrites bound antibodies within the hippocampus and the lateral septum, whereas both dendrites and neuronal cell bodies were immunoreactive in the medial septum, in the diagonal band of Broca, and in the dorsal and median raphe nuclei. Therefore, differential addressing of serotonin1A receptors could occur from one neuron to another. In general, the distribution and density of serotonin1A receptor-like immunoreactivity in the whole brain and in spinal cord were consistent with the mapping of serotonin1A receptor binding sites and serotonin1A receptor mRNA previously established by immunoautoradiographic and in situ hybridization procedures.

Antibodies and antisense oligonucleotide for probing the distribution and putative functions of central 5-HT6 receptors.

Among the recently cloned serotonin (5-hydroxytryptamine, 5-HT) receptors, the 5-HT6 subtype is of special interest for at least two reasons: 1) it is abundant in limbic areas which participate in the control of mood and emotion; and 2) some antidepressants and antipsychotics are potent 5-HT6 receptor antagonists. Studies using polyclonal anti-5-HT6 receptor antibodies and an antisense oligonucleotide were performed in order to investigate further the function(s) of 5-HT6 receptors in the rat brain. Immunocytochemistry at the light and electron microscope levels showed that 5-HT6 receptors are mainly confined to the dendritic compartment, suggesting that they could mediate 5-HT actions on neuronal firing. In some limbic areas, 5-HT6 receptor-like immunoreactivity is also associated with neuronal cilia with yet unknown functions. Continuous i.c.v. infusion with an antisense oligonucleotide for 3-4 days resulted in decreased 5-HT6 receptor-like immunostaining of the nucleus accumbens and anxiogenic behaviours in the social interaction and elevated plus maze tests. Selective 5-HT6 receptor ligands are eagerly expected to investigate further the potential implication of these receptors in limbic-dependent behaviours.

Quantification of 5-hydroxytryptamine1A receptors in the cerebellum of normal and X-irradiated rats during postnatal development.

5-Hydroxytryptamine1A receptors were studied in rats during the first postnatal month in the normal cerebellum and in the granule cell-deprived cerebellum produced by X-irradiation at postnatal day 5. Quantitative autoradiographic studies on sagittal sections of cerebellar vermis, using [1251]BH-8-MeO-N-PAT as radioligand or specific anti-receptor antibodies, revealed that 5-hydroxytryptamine1A receptors existed in the molecular/Purkinje cell layer but at variable density from one lobule to another. Thus, in both normal and X-irradiated rats, the posterior lobules were more heavily labelled than the anterior ones, and the density of 5-hydroxytryptamine1A sites decreased progressively in all the cerebellar folia down to hardly detectable levels at postnatal day 21. However, the intensity of labelling remained higher at postnatal day 8 and postnatal day 12 in X-irradiated rats than in age-paired controls. Measurements of [3H]8-OH-DPAT specific binding to membranes from whole cerebellum confirmed that the density of 5-hydroxytryptamine1A sites per mg membrane protein (Bmax) was higher in X-irradiated animals than in age-paired controls. However, on a "per cerebellum" basis, no significant difference could be detected between the total number of 5-hydroxytryptamine1A sites, which progressively increased in both control and X-irradiated animals during the first postnatal month. These results therefore show that 5-hydroxytryptamine1A receptors are not located on developing granule cells. The progressive decrease in 5-hydroxytryptamine1A receptor density during the first postnatal month did not reflect a transient expression of 5-hydroxytryptamine1A receptors in the cerebellum of newborn rats, but resulted from the progressive "dilution" of these sites in this growing structure. The higher density of 5-hydroxytryptamine1A sites in X-irradiated rats simply reflected a lower "dilution" due to the delayed growth of the cerebellum in these animals.

Selective irreversible blockade of 5-hydroxytryptamine1A and 5-hydroxytryptamine1C receptor binding sites in the rat brain by 8-MeO-2'-chloro-PAT: a quantitative autoradiographic study.

The possible irreversible blockade of 5-hydroxytryptamine1 receptor subtypes 5-hydroxytryptamine1A, 5-hydroxytryptamine1B/5-hydroxytryptamine1D and 5-hydroxytryptamine1C by the chloramine 8-methoxy-2-(N-2'-chloropropyl,N-propyl)aminotetralin (8-MeO-2'-chloro-PAT) was investigated in rat brain sections by quantitative autoradiography using [3H]8-hydroxy-2-(di-n-propylamino)tetralin [( 3H]8-OH-DPAT), [3H]5-hydroxytryptamine, [125I]BH-8-MeO-N-PAT and [125I]cyanopindolol as radio-ligands. A marked reduction (-50% to -75%) of [3H]8-OH-DPAT and [125I]BH-8-MeO-N-PAT specific binding to 5-hydroxytryptamine1A sites in the hippocampus (CA1 area) and the dorsal raphe nucleus, and of [3H]5-hydroxytryptamine specific binding to 5-hydroxytryptamine1C sites in the choroid plexus was found in sections exposed to 1 microM 8-MeO-2'-chloro-PAT and then washed extensively. In contrast the specific binding of [3H]5-hydroxytryptamine to 5-hydroxytryptamine1B/5-hydroxytryptamine1D sites and of [125I]cyanopindolol to 5-hydroxytryptamine1B sites in the substantia nigra and dorsal subiculum remained unaltered by this treatment. Similarly [125I]cyanopindolol binding to beta-adrenergic receptors was not affected by 8-MeO-2'-chloro-PAT. Prior occupancy of 5-hydroxytryptamine1A sites by 10 microM 5-hydroxytryptamine or 8-OH-DPAT, and of 5-hydroxytryptamine1C sites by 10 microM 5-hydroxytryptamine prevented any subsequent blockade by 8-MeO-2'-chloro-PAT. These data indicate that 8-MeO-2'-chloro-PAT should be a useful alkylating agent for achieving selective irreversible blockade of central 5-hydroxytryptamine1A and 5-hydroxytryptamine1C receptors in vivo in the rat.

Immunolabeling of the rat central nervous system with antibodies partially selective of the short form of the 5-HT3 receptor.

Polyclonal antibodies were raised against a synthetic hexadecapeptide corresponding to the portion of the second intracytoplasmic loop of the short form of the mouse 5-hydroxytryptamine-3A receptor subunit (5-HT3A-S), which differs from the long form (5-HT3A-L) by the removal of six amino acids. Antibodies were detected by enzyme-linked immunosorbent assay as soon as two months after the first injection to rabbits of the peptide coupled to keyhole limpet hemocyanin. Immunoblot detection of fusion proteins comprising glutathione-S-transferase and the second intracellular loop of 5-HT3A-S or 5-HT3A-L, and immunoprecipitation of cloned receptors showed that antibodies exhibited some selectivity for the short variant. Affinity chromatography allowed the purification of selective anti-5-HT3A-S antibodies which yielded a strong positive labeling of plasma membrane, reticulum and Golgi apparatus of COS-7 cells expressing murine 5-HT3A-S. In contrast, COS-7 cells expressing similar levels of 5-HT3A-L exhibited only a very weak labeling. Selectivity was also observed on immunoblots of cloned receptors transiently expressed in COS-7 cells, or stably expressed in CHO cells, both systems showing an immunolabeled component at 53,000-54,000 mol. wt. Immunoautoradiographic labeling of central nervous system sections showed that 5-HT3A-S-like immunoreactivity was found mostly within the nucleus of the solitary tract, the nucleus of the spinal tract of the trigeminal nerve, and the dorsal horn of the the spinal cord in the rat. After unilateral ablation of the nodose ganglion, 5-HT3A-S-like immunoreactivity decreased markedly in the ipsilateral part of the nucleus of the solitary tract, as expected of the presynaptic localization of 5-HT3 receptors. Finally, immunohistochemistry at the light and electron microscope levels revealed that 5-HT3A-S-like immunoreactivity was associated essentially with terminals and axonal profiles. All these results demonstrate that the immunolabeling exhibited by these antibodies is consistent with a specific and partially selective recognition of the short isoform of the 5-HT3A subunit. Because the pattern of immunoautoradiographic labeling matches the distribution previously established with selective radioligands, it can be inferred that these antibodies probably recognized the same fully assembled form of the 5-HT3A-S receptor subunit.

Serotonin1A receptors are expressed by a subpopulation of cholinergic neurons in the rat medial septum and diagonal band of Broca--a double immunocytochemical study.

The possible colocalization of 5-hydroxytryptamine1A receptors and choline acetyltransferase in the same neurons of the medial septum and diagonal band of Broca was investigated using double immunocytochemical techniques, either on the same section or on adjacent thin sections of the rat brain. The presence of both antigens in the same neurons was demonstrated at the light and electron microscopic levels. The proportion of cholinergic neurons that express 5-hydroxytryptamine1A receptors was similar in the different parts of the septal complex (around 25%). By contrast, the proportion of 5-hydroxytryptamine1A receptor-positive neurons also exhibiting choline acetyltransferase immunoreactivity was much higher (40-44%) in the dorsal and ventral groups of cholinergic cells, than in the intermediate group (18%). In line with the topographical distribution of cholinergic projections, this result points out the potential involvement of 5-hydroxytryptamine1A receptors in the control of the septohippocampal cholinergic projection by serotonin. This connection might be relevant to learning and memory, and in the appearance of age-dependent or neurodegenerative cognitive deficits, which have been shown to involve alterations in both the serotoninergic and the cholinergic systems.

Cellular and subcellular localization of 5-hydroxytryptamine1B receptors in the rat central nervous system: immunocytochemical, autoradiographic and lesion studies.

The localization of 5-hydroxytryptamine1B receptors in the rat central nervous system was investigated using anti-peptide antibodies that recognize a selective portion of the third intracytoplasmic loop of the receptor protein. At the light microscope level the densest 5-hydroxytryptamine1B receptor-like immunoreactivity was observed in ventral pallidum, globus pallidus, substantia nigra and dorsal subiculum. In addition, moderate immunoreactivity was found in the entopeduncular nucleus, the superficial gray layer of the superior colliculus, the caudate-putamen and the deep nuclei of the cerebellum. This distribution matched perfectly that previously described from radioligand binding studies. At the ultrastructural level, 5-hydroxytryptamine1B receptor-like immunoreactivity was associated with axons and axon terminals in the three areas examined: substantia nigra, globus pallidus and superficial gray layer of the superior colliculus. In all cases, immunostaining was located on the plasma membrane of unmyelinated axon terminals and in the cytoplasm close to the plasmalemma. Synaptic differentiations were never labelled but, in some cases, 5-hydroxytryptamine1B receptor-like immunoreactivity was found in their close vicinity. Injection of kainic acid into the neostriatum resulted in a marked decrease in receptor-like immunoreactivity in the globus pallidus and the substantia nigra, consistent with the location of 5-hydroxytryptamine1B receptors on terminals of striatopallidal and striatonigral fibres, respectively. A reduction in 5-hydroxytryptamine1B receptor-like immunoreactivity was also noted in the superficial gray layer of the superior colliculus after contralateral enucleation, as expected of the location of 5-hydroxytryptamine1B receptors on the terminals of retinocollicular fibres. In both lesion experiments, immunolabelled degenerating terminals were observed in the projection areas. Anterograde labelling experiments coupled with immunocytochemical detection further showed that 5-hydroxytryptamine1B receptors in the substantia nigra are located on axons of striatal neurons. These data provide anatomical support for the idea that 5-hydroxytryptamine1B receptors act as terminal receptors involved in presynaptic regulation of the release of various neurotransmitters, including 5-hydroxytryptamine itself.