Methods for analysis of brain connectivity: An IFCN-sponsored review.

The goal of this paper is to examine existing methods to study the "Human Brain Connectome" with a specific focus on the neurophysiological ones. In recent years, a new approach has been developed to evaluate the anatomical and functional organization of the human brain: the aim of this promising multimodality effort is to identify and classify neuronal networks with a number of neurobiologically meaningful and easily computable measures to create its connectome. By defining anatomical and functional connections of brain regions on the same map through an integrated approach, comprising both modern neurophysiological and neuroimaging (i.e. flow/metabolic) brain-mapping techniques, network analysis becomes a powerful tool for exploring structural-functional connectivity mechanisms and for revealing etiological relationships that link connectivity abnormalities to neuropsychiatric disorders. Following a recent IFCN-endorsed meeting, a panel of international experts was selected to produce this current state-of-art document, which covers the available knowledge on anatomical and functional connectivity, including the most commonly used structural and functional MRI, EEG, MEG and non-invasive brain stimulation techniques and measures of local and global brain connectivity.

The solitary chemosensory cells and the diffuse chemosensory system of the airway.

Solitary chemosensory cells (SCCs), which resemble taste bud cells, are present in the epidermis and oropharynx of most primary aquatic vertebrates. Recent studies have led to the description of SCCs also in mammals too. In the airway and digestive apparatus, these elements form a diffuse chemosensory system. SCCs do not aggregate into groups and in SCCs, as in taste bud cells, immunoreactivity forthe G-protein subunit alpha-gustducin and for other molecules of the chemoreceptive cascade was found. Questions remain about the role of the diffuse chemosensory system in control of complex functions (e.g. airway surface liquid secretion) and about the involvement of chemoreceptors in respiratory diseases. Therapeutic actions targeting chemoreceptors could be tested in the treatment of respiratory diseases.

1898: the Golgi apparatus emerges from nerve cells.

A century ago, Camillo Golgi discovered in neurons an intracellular network of anastomosing threads, impregnated by the chromoargentic reaction he had devised to stain the nervous tissue. This structure, designated by Golgi as 'internal reticular apparatus', was soon detected in a wide variety of eukaryotic cells. However, skepticism arose on the existence of the Golgi apparatus in the first decades of this century, when it was fiercely debated whether this structure represented a genuine new cell constituent or an artifact due to the deposit of metallic impregnation on diverse cytoplasmic structures. The reality of the Golgi apparatus became established unequivocally only with the application of electron microscopy; with the visualization of its fine structure, the apparatus finally achieved the status of cytoplasmic organelle, and thus, linked with Golgi's name, entered the modern era of investigation on its components, chemistry and function.

Trypanosoma brucei and the nervous system.

African sleeping sickness, characterized by a peculiar pain syndrome and prominent neuropsychiatric symptoms, is caused by the parasite Trypanosoma brucei (T.b.). In experimental T.b. infections, a molecule released from the trypanosomes has been isolated that binds to the CD8 molecule of T cells, whereby T cells are activated to secrete interferon gamma. This cytokine binds to the parasites and triggers them to proliferate, establishing a peculiar bidirectional activating signal system. The hypothesis is presented that the molecules involved in these bidirectional signals might also interact with neurons, thus causing brain dysfunctions. Studies on the molecular interactions between parasites and the nervous system in sleeping sickness might reveal basic mechanisms underlying other neuropsychiatric diseases.

Cytokine-induced activation of glial cells in the mouse brain is enhanced at an advanced age.

Numerous neurological diseases which include neuroinflammatory components exhibit an age-related prevalence. The aging process is characterized by an increase of inflammatory mediators both systemically and in the brain, which may prime glial cells. However, little information is available on age-related changes in the glial response of the healthy aging brain to an inflammatory challenge. This problem was here examined using a mixture of the proinflammatory cytokines interferon-gamma and tumor necrosis factor-alpha, which was injected intracerebroventricularly in young (2-3.5 months), middle-aged (10-11 months) and aged (18-21 months) mice. Vehicle (phosphate-buffered saline) was used as control. After a survival of 1 or 2 days (all age groups) or 4 days (young and middle-aged animals), immunohistochemically labeled astrocytes and microglia were investigated both qualitatively and quantitatively. In all age groups, astrocytes were markedly activated in periventricular as well as in deeper brain regions 2 days following cytokine treatment, whereas microglia activation was already evident at 24 h. Interestingly, cytokine-induced activation of both astrocytes and microglia was significantly more marked in the brain of aged animals, in which it included numerous ameboid microglia, than of younger age groups. Moderate astrocytic activation was also seen in the hippocampal CA1 field of vehicle-treated aged mice. FluoroJade B histochemistry and the terminal deoxynucleotidyl transferase-mediated UTP nick-end labeling technique, performed at 2 days after cytokine administration, did not reveal ongoing cell death phenomena in young or aged animals. This indicated that glial cell changes were not secondary to neuronal death. Altogether, the findings demonstrate for the first time enhanced activation of glial cells in the old brain, compared with young and middle-aged subjects, in response to cytokine exposure. Interestingly, the results also suggest that such enhancement does not develop gradually since youth, but appears characterized by relatively late onset.

Drug resistance and hippocampal damage after delayed treatment of pilocarpine-induced epilepsy in the rat.

Temporal lobe epilepsy (TLE) is the most common and pharmacoresistant form of epilepsy. Problems that cause pharmacoresistance may include delayed therapy due to late consultation, especially in developing countries. Our study aimed at unraveling consequences of delayed drug treatment using a rat model of TLE. Following pilocarpine-induced status epilepticus interrupted after 4h, rats were continuously videorecorded for onset and recurrence of spontaneous convulsive seizures. The animals were then treated for 50 days with carbamazepine (CBZ; first-line drug in TLE and effective also in rats), starting at seizure onset (27.22+/-3.38 days after status epilepticus) or 50 days later, and compared with epileptic untreated rats and non-epileptic CBZ-treated ones. Convulsive seizure frequency and duration, and hippocampal cell changes were evaluated. In particular, parvalbumin-containing hippocampal interneurons, astrocytes and microglia were characterized with immunohistochemistry and quantitative analyses. Prompt administration of CBZ suppressed seizures; delayed treatment only decreased frequency of convulsive seizures, which were also relatively prolonged. In hippocampal regions, histopathological damage, parvalbumin immunoreactivity loss, and glial activation were very marked after delayed treatment, and were reduced only slightly compared to untreated epilepsy, but enhanced compared to early treatment. The data on high frequency and duration of convulsive seizures in late-therapy rats indicate that delayed CBZ administration caused a high degree of drug resistance. This condition was subserved by severe damage in the hippocampus, presumably consequent to long-term seizure recurrence. Overall the data indicate that the paradigm of delayed treatment of limbic epilepsy could provide a model of drug-refractory TLE with hippocampal sclerosis.

Low-dose dipyridamole infusion acutely increases exercise capacity in angina pectoris: a double-blind, placebo controlled crossover stress echocardiographic study.

OBJECTIVES: The aim of this study was to assess whether endogenous accumulation of adenosine, induced by low-dose dipyridamole infusion, protects from exercise-induced ischemia. BACKGROUND: Adenosine is a recognized mediator of ischemic preconditioning in experimental settings. METHODS: Ten patients (all men: mean age 63.4 +/- 7.3 years) with chronic stable angina, angiographically assessed coronary artery disease (n = 7) or previous myocardial infarction (n = 3) and exercise-induced ischemia underwent on different days two exercise-stress echo tests after premedication with placebo or dipyridamole (15 mg in 30 min, stopped 5 min before testing) in a double-blind, placebo controlled, randomized crossover design. RESULTS: In comparison with placebo, dipyridamole less frequently induced chest pain (20% vs. 100%, p = 0.001) and >0.1 mV ST segment depression (50% vs. 100%, p < 0.05). Wall motion abnormalities during exercise-stress test were less frequent (placebo = 100% vs. dipyridamole = 70%, p = ns) and significantly less severe (wall motion score index at peak stress: placebo = 1.55 +/- 0.17 vs. dipyridamole = 1.27 +/- 0.2, p < 0.01) following dipyridamole, which also determined an increase in exercise time up to echocardiographic positivity (placebo = 385.9 +/- 51.4 vs. dipyridamole = 594.4 +/- 156.9 s, p < 0.01). CONCLUSIONS: Low-dose dipyridamole infusion increases exercise tolerance in chronic stable angina, possibly by endogenous adenosine accumulation acting on high affinity A1 myocardial receptors involved in preconditioning or positively modulating coronary flow through collaterals.

Graves' disease and thyroxine-binding globulin deficiency.

Thyroxine-binding globulin (TBG) deficiency has been frequently described in single patients and in many families. Most people with abnormal TBG concentrations are euthyroid. Cases of Graves' disease and TBG deficit have rarely been reported. We describe the case of a person with Graves' disease and TBG deficiency. Because of this condition, the patient had a misdiagnosis during part of his clinical history, and therefore underwent unnecessary therapy.

The effect of diltiazem, a calcium channel-blocking drug, on cardiac rate and rhythm in hyperthyroid patients.

Tachycardia and tachyarrhythmias are frequent in patients with thyrotoxicosis, especially in the elderly. Since myocardial calcium uptake is increased in thyrotoxic rats, the efficacy of the calcium channel-blocking drug diltiazem in decreasing heart rate and the incidence of arrhythmias was evaluated in 11 hyperthyroid patients. All patients were studied with a 24-hour Holter monitor prior to the beginning of sole diltiazem therapy (120 mg given every eight hours), on the tenth day of therapy, and five days after therapy was discontinued. Heart rate significantly decreased by 17% during diltiazem treatment (96.5 +/- 3.7 systoles/min vs 79.9 +/- 3.2 systoles/min [mean +/- SE]) and returned to baseline values five days after the therapy was discontinued (100.7 +/- 3.4 systoles/min). Similarly, the number of premature ventricular extrasystoles per hour was significantly decreased (18 +/- 7 vs 2 +/- 1). In three patients, asymptomatic bouts of supraventricular tachycardia, paroxysmal atrial fibrillation, or ventricular tachycardia disappeared during diltiazem therapy. These findings suggest that calcium-blocking drugs may be extremely useful as adjunctive therapy for thyrotoxicosis in the presence of angina, congestive failure, and tachyarrhythmias.

Alterations of orexinergic and melanin-concentrating hormone neurons in experimental sleeping sickness.

Human African trypanosomiasis or sleeping sickness is a severe, neglected tropical disease caused by the extracellular parasite Trypanosoma brucei. The disease, which leads to chronic neuroinflammation, is characterized by sleep and wake disturbances, documented also in rodent models. In rats and mice infected with Trypanosoma brucei brucei, we here tested the hypothesis that the disease could target neurons of the lateral hypothalamus (LH) containing orexin (OX)-A or melanin-concentrating hormone (MCH), implicated in sleep/wake regulation. In the cerebrospinal fluid of infected rats, the OX-A level was significantly decreased early after parasite neuroinvasion, and returned to the control level at an advanced disease stage. The number of immunohistochemically characterized OX-A and MCH neurons decreased significantly in infected rats during disease progression and in infected mice at an advanced disease stage. A marked reduction of the complexity of dendritic arborizations of OX-A neurons was documented in infected mice. The evaluation of NeuN-immunoreactive neurons did not reveal significant neuronal loss in the LH of infected mice, thus suggesting a potential selective vulnerability of OX-A and MCH neurons. Immunophenotyping and quantitative analysis showed in infected mice marked activation of microglial cells surrounding OX-A neurons. Day/night oscillation of c-Fos baseline expression was used as marker of OX-A neuron activity in mice. In control animals Fos was expressed in a higher proportion of OX-A neurons in the night (activity) phase than in the day (rest) phase. Interestingly, in infected mice the diurnal spontaneous Fos oscillation was reversed, with a proportion of OX-A/Fos neurons significantly higher at daytime than at nighttime. Altogether the findings reveal a progressive decrease of OX-A and MCH neurons and dysregulation of OX-A neuron diurnal activity in rodent models of sleeping sickness. The data point to the involvement of these peptidergic neurons in the pathogenesis of sleep/wake alterations in the disease and to their vulnerability to inflammatory signaling.

Altered neuronal activity rhythm and glutamate receptor expression in the suprachiasmatic nuclei of Trypanosoma brucei-infected rats.

The parasites Trypanosoma brucei cause African trypanosomiasis (sleeping sickness), a severe neuropsychiatric disease with marked disturbances of sleep-wake alternation. The sites of brain lesions are not well characterized. The present experimental investigation is focused on the hypothalamic suprachiasmatic nuclei, which play a role of a biological clock entraining endogenous rhythms in the mammalian brain. The electrophysiological properties of these neurons were analyzed in slice preparations from trypanosome-infected rats. The neuronal spontaneous activity, which shows a circadian oscillation, was markedly altered in the infected animals, displaying a reduced firing rate and phase advance of its circadian peak. The direct retinal fibers, which play a pivotal role in entrainment of the circadian pacemaker, displayed a normal density and distribution in the suprachiasmatic nuclei of infected animals after intraocular tracer injections in vivo. At the postsynaptic level, immunohistochemistry and Western blotting revealed in the suprachiasmatic nuclei of infected rats a selective decrease of the expression of glutamate AMPA GluR2/3 and NMDAR1 receptor subunits that gate retinal afferents. These data disclose an impairment of the neuronal functions in the biological clock in African trypanosomiasis, and may serve to unravel functional and molecular mechanisms behind endogenous rhythm disturbances.

Diencephalic asymmetries.

Structural asymmetry in diencephalic regions has been reported in a number of studies since the pioneering observations by Kemali and Braitenberg, Atlas of the frog's brain. Springer Verlag: 1969. Anatomical differences between the left and right habenulae have been identified in many lower vertebrate species. While there are few reports of structural asymmetry in the dorsal thalamus, there is evidence that asymmetrical thalamofugal projections can be induced in the visual system of chicks by lateralized sensory stimulation prior to hatching. Finally, there have been consistent reports of differences between and right sides of the hypothalamus in their sensitivity to the effects of circulating gonadal hormones in rats. In most cases, these asymmetries are sex-linked and correspond to a lateralization of function. Although the significance of these diencephalic asymmetries is still enigmatic, their existence indicates that asymmetry is not a phylogenetically recent feature of the brain, and the left-right differences in the brain may be mediated by a common ontogenetic mechanism and may underlie the development of highly specialized functions.

Echocardiographic left ventricular hypertrophy as related to arterial pressure and plasma norepinephrine concentration in arterial hypertension. Reversal by atenolol treatment.

We tried to assess relationships between echocardiographic left ventricular hypertrophy (LVH), arterial pressure levels, and plasma norepinephrine concentration (NE) in 20 previously untreated stable hypertensive patients with LVH, and in 11 healthy normotensive control subjects. Interventricular septal (IVS) thickness, posterior wall (PW) thickness, and left ventricular mass index (LVMI) were related to arterial pressure levels and to NE by univariate and multivariate regression analyses. In addition, after 18 months of monotherapy with atenolol (carried out in nine of 20 patients), the relationship between echocardiographic changes and degree of pressure reduction was tested. Before treatment, PW thickness weakly correlated with systolic (r = 0.55; p less than 0.01) and mean (r = 0.50; p less than 0.05) arterial pressure. IVS thickness weakly correlated with NE (r = 0.53; p less than 0.05). On this relatively small sample, multivariate regression analysis showed an association of both IVS thickness (R = 0.57; p less than 0.05) and PW thickness (R = 0.58; p less than 0.05) with mean arterial pressure (MAP) and NE. After atenolol, there was a reduction in IVS thickness (1.15 to 1.02 cm; p less than 0.01), PW thickness (1.08 to 0.99 cm; p less than 0.01), and LVMI (136.3 to 113.8 g/m2; p less than 0.01), besides a significant reduction in blood pressure and heart rate. The degree of pressure reduction induced by treatment did not correlate the change in IVS or PW thickness. In contrast, the change in diastolic and mean arterial pressure positively correlated the change in LVMI (r = 0.72 and r = 0.75, respectively; both p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Low-dose captopril therapy in mild and moderate hypertension. Randomized comparison of twice daily vs three times daily doses.

We have investigated the antihypertensive activity of relatively low daily doses of captopril in patients with mild and moderate arterial hypertension. In a first trial, at the end of a 2-week placebo washout period, 18 patients with essential hypertension WHO Stage I or II were treated with captopril, 25 mg three times daily (t.i.d.), 25 mg twice daily (b.i.d.), 50 mg t.i.d., and 50 mg b.i.d., according to a randomized within-patient open design, with each regimen lasting for a 2-week period. In a second trial, 12 hypertensive patients not adequately controlled by chlorthalidone 25 mg daily as monotherapy (supine diastolic blood pressure at rest greater than 95 mm Hg), continued the diuretic treatment in combination with captopril, 25 mg t.i.d. and 25 mg b.i.d. according to a randomized within-patient open design. Analysis of variance did not reveal differences between the four captopril dosing schedules (1st trial), or between the two captopril dosing schedules (2nd trial). Both the patients on captopril monotherapy (1st trial) and those cotreated with chlorthalidone (2nd trial) showed lower systolic and diastolic blood pressure values on each captopril regimen compared to prerandomization values (all p less than 0.01). No relevant unwanted effects were noted. We conclude that in patients with mild or moderate essential hypertension, either untreated or resistant to chlorthalidone, captopril is effective in reducing blood pressure even at daily doses not exceeding 150 mg, without differences between a t.i.d. and a b.i.d. dosing schedule.

Noninvasive assessment of chronotropic and inotropic response to preferential beta-1 and beta-2 adrenoceptor stimulation.

The relative chronotropic and inotropic activity of preferential beta 1- and beta 2-adrenoceptor stimulation was investigated in seven healthy male subjects in a randomized within-subject, single-blind study. Two doses of beta 1-selective agonist prenalterol (1 mg/hr or 2 mg/hr) and of beta 2-selective agonist salbutamol (300 micrograms/hr or 600 micrograms/hr) were infused intravenously in four separate sessions, with intervals of at least 48 hr between sessions. At each session cuff blood pressure and heart rate (HR) were measured and some hemodynamic information on the inotropic state were derived by echocardiography. Both prenalterol and salbutamol induced increases in HR, but tachycardia was greater after salbutamol, whereas the positive inotropic response to beta-stimulation was greater after prenalterol. At comparable HR rises (prenalterol, from 66.0 +/- 5.5 to 72.2 +/- 4 bpm; salbutamol, from 64.6 +/- 6 to 70.0 +/- 7 bpm), inotropic response seemed to be greater after prenalterol than after salbutamol (systolic blood pressure [SBP]: 133.5 +/- 8 and 120.7 +/- 8 mm Hg; mean velocity of circumferential fiber shortening [Vcf]: 1.54 +/- 0.13 and 1.31 +/- 0.12 c/s; ejection fraction [EF]: 72.4% +/- 5% and 69.5% +/- 4%; stroke index: 47.4 +/- 4 and 41.7 +/- 3 ml/m2). In presence of a chronotropic effect (HR from 64.6 +/- 6 to 70.0 +/- 7 bpm), the low salbutamol dose did not induce any changes in the indices of inotropism (SBP: from 119.2 +/- 6 to 120.7 +/- 8 mm Hg; mean Vcf: from 1.28 +/- 0.11 to 1.31 +/- 0.12 c/s; EF: from 68.1% +/- 5% to 69.5% +/- 4%; stroke index: from 40.2 +/- 3 to 41.7 +/- 3 ml/m2.(ABSTRACT TRUNCATED AT 250 WORDS)

Nerve Growth Factor Receptor-immunoreactive Fibres Innervate the Reticular Thalamic Nucleus: Modulation by Nerve Growth Factor Treatment in Neonate, Adult and Aged Rats.

Terminal arborizations expressing nerve growth factor receptor (NGF-R) have been detected with immunohistochemistry in the reticular thalamic nucleus of neonate, adult and aged rats. Intracerebroventricular administration of nerve growth factor (NGF) resulted in a dramatic increase in NGF-R immunoreactivity throughout the lifespan. This effect was paralleled by a concomitant increase in NGF-R immunopositivity in the neurons of the basal forebrain, which was here demonstrated also in aged animals, thus indicating that the NGF-R immunoreactivity within the reticular thalamic nucleus derives in all likelihood from cholinergic neuronal cell bodies of the basal forebrain. Our results demonstrate a prominent ability of NGF to up-regulate its receptors within fibres innervating the reticular thalamic nucleus, and show that this up-regulation of NGF-R is maintained throughout the lifetime. Altogether this indicates that the reticular thalamic nucleus may represent a new, important site of action of endogenous NGF or NGF-like molecules within the brain. In view of the crucial role played by the reticular thalamic nucleus in gating thalamocortical information, the autoregulation of NGF-R in this structure may have important concomitants in both physiological and pathological conditions.

Crosstalk between the two sides of the thalamus through the reticular nucleus: a retrograde and anterograde tracing study in the rat.

In order to investigate the possible routes linking the thalamus in the two sides of the brain, the connections of the reticular nucleus (RT), the major component of the ventral thalamus, with contralateral dorsal thalamic nuclei were systematically investigated in the adult rat. This study was performed with several tract-tracing techniques: single and double retrograde labeling with fluorescent tracers, and anterograde tracing with biocytin. Retrograde tracing was also combined with immunocytochemistry to provide additional criteria for the identification of labeled RT neurons. The data obtained with the retrograde transport of one fluorescent tracer showed that RT neurons project to contralateral dorsal thalamic domains. In particular, retrograde labeling findings indicated that the anterior intralaminar nuclei, as well as the ventromedial (VM) nucleus, are preferential targets of the contralateral RT projections. Commissural neurons were concentrated in two portions of RT: its rostral part, including the rostral pole, which projects to the contralateral central lateral (CL) and paracentral (Pc) nuclei, and the ventromedial sector of the middle third of RT, which projects to the contralateral VM and posterior part of CL and Pc. The double retrograde labeling study of the bilateral RT-intralaminar connection indicated that at least part of the commissural RT cells bifurcate bilaterally to symmetrical portions of the anterior intralaminar nuclei. The targets of the RT commissural system inferred from the retrograde labeling data were largely confirmed by anterograde tracing. Moreover, it was shown that RT fibers cross the midline in the intrathalamic commissure. The present data demonstrate that bilateral RT connections with the dorsal thalamus provide a channel for interthalamic crosstalk. Through these bilateral connections with thalamic VM and intralaminar neurons, RT could influence the activity of wide territories of the cerebral cortex and basal ganglia of both hemispheres.

Corticospinal neurons with branching axons to the dorsal column nuclei in the monkey.

Previous work in cats has shown that cells of origin of the corticospinal tract give rise to collateral branches to the dorsal column nuclei (DCN). The present experiments were performed in monkeys (Macaca fascicularis) in which 2% fast blue and 2% diamidino yellow were delivered to infiltrate the dorsolateral funiculus at levels between C2 and C6 and the cuneate nucleus on the same side. Retrograde labelling in the cortex allows simultaneous visualization of three classes of neurons: corticospinal tract (CST) neurons, corticocuneate tract (CCT) neurons, and double-labelled neurons. The morphological features and distribution of CST and CCT neurons are similar to those previously reported from investigations based mainly upon the retrograde transport of horseradish peroxidase (HRP). CST neurons occur in layer V in the pre- and postcentral gyri, except for the lateral part (face representation), in the supplementary motor and sensory cortex, and in SII. CCT neurons are present in layer V largely in the postcentral gyrus and in SII. Double-labelled neurons are present wherever CST and CCT neurons are found. Reconstruction and quantitative data from the pericentral cortex show that up to 60% of CCT neurons are double-labelled and are found predominantly in areas 1 and 2, and that their perikarya are in the size range of the larger CCT neurons. Comparison of these results with those obtained previously in cats by using HRP and tritiated, enzymatically inactive HRP (3H-apo-HRP, Rustioni and Hayes: Exp. Brain Res. 43:237-245, 1981) suggests that CST neurons with branching axons to the DCN are considerably more numerous in monkeys than in cats. To determine whether this difference is caused by the different tracers used in the two species. 2% fast blue and 2% diamidino yellow were delivered in cats to infiltrate the dorsolateral funiculus at C2-C3 and the cuneate nucleus on the same side. The results in these cats are remarkably similar to those obtained in the previous study, which used HRP and 3H-apo-HRP: double-labelled neurons occur predominantly in area 3a and constitute 14-16% of the CCT neurons in the pericruciate area. The results bear upon mechanisms of descending control and tuning of performances that characterize the dorsal column-medial lemniscal system, e.g., discrimination of discrete spatiotemporal cues. The species differences may be related to the higher degree of tactile resolution and synchronous control of sensory inflow at the DCN and spinal cord in monkeys relative to cats.

Degranulation, density, and distribution of mast cells in the rat thalamus: a light and electron microscopic study in basal conditions and after intracerebroventricular administration of nerve growth factor.

In the adult rat brain mast cells reside selectively in the thalamus. We investigated thalamic mast cells stained by acidic toluidine blue or pinacyanol, and with histamine immunocytochemistry, focusing on their state of activity revealed by degranulation. Mast cells exhibited perivascular prevalence and high quantitative variability, between cases and in different sections, with no asymmetry or topographical selectivity in thalamic nuclei. Pinacyanol, alone or with erythrosine, stained mast cells with higher sensitivity than toluidine blue. However, toluidine blue was highly predictive of pinacyanol staining and provided the best resolution of mast cell cytoplasmic features. Histamine immunocytochemistry labeled 61% of pinacyanol-stained mast cells. Intensely toluidine blue-stained granulated cells, as well as cells exhibiting different degrees of degranulation that paralleled lighter staining, were observed. The response of thalamic mast cells to intracerebroventricular administration of nerve growth factor (NGF) and control cytochrome-c injections was evaluated after 2, 24, and 72 hours. No obvious changes in mast cell number or distribution were found after treatment, but massive degranulation was frequently observed after NGF administration. Significant decrease of staining intensity of mast cells, supporting enhanced degranulation, was documented in NGF-treated animals by quantitative image analysis. Ultrastructural features of mast cell degranulation, with granule coalescence and matrix dissolution, were detected in untreated and NGF-treated cases. The findings point out that mast cells are active in the thalamus in basal conditions and that NGF has the potential to elicit long-lasting degranulation of thalamic mast cells in vivo, exerting a direct effect and/or priming these cells to react to endogenous stimuli.

Nitric oxide synthase in the adult and developing thalamus: histochemical and immunohistochemical study in the rat.

The distribution of neuronal elements that express nitric oxide synthase (NOS), the synthetic enzyme of the free radical nitric oxide, was investigated in the adult and developing rat thalamus by means of NADPH-diaphorase (NADPH-d) histochemistry, which is a marker of NOS. Immunocytochemistry was also used to confirm the equivalence between the histochemical pattern of staining and the distribution of the expression of the neuronal NOS isoform. In the adult thalamus, NADPH-d-positive and NOS-immunoreactive perikarya were selectively concentrated along the midline (in the paraventricular, rhomboid, and central medial nuclei) and in the dorsal and ventral lateral geniculate nuclei. Isolated clusters of stained neurons were also observed in the lateral posterior nucleus, in the dorsal part of the medial geniculate nucleus, and in the ventromedial nucleus. Positive perikarya were either absent or very sparse in the other thalamic nuclei. Many thalamic domains were, however, characterized by distinct patterns of NADPH-d-positive fibers, preterminal and terminal-like elements. The highest density of stained neuropil was observed in the anteroventral and anteromedial nuclei, in several of the midline nuclei, in the anterior intralaminar nuclei, and in the lateral and medial geniculate nuclei. Although histochemical reactivity was observed in the thalamus at birth, the intensity and the pattern of distribution of staining observed in adulthood was not achieved until the end of the third postnatal week. The NADPH-d histochemical positivity followed discrete developmental schedules in various thalamic domains, and different areas reached a mature pattern at different ages. In addition, populations of transiently stained neuronal cell bodies were observed in the medial thalamus during the first two postnatal weeks. These results show discrete patterns of expression of NOS in the adult and developing thalamus and suggest that nitric oxide may be involved in selected physiological and developmental roles in different thalamic domains.