Italian Study on Depressive Disorders (STudio Italiano MAlattia Depressiva, or STIMA-D): a nationwide snapshot of the status of treatment for major depression.

INTRODUCTION: Major depression is a worldwide problem and often remains undetected and untreated. Given the low rates of detection plus the need to intervene in a short time, it is important to identify factors which are likely to improve treatment outcomes. METHODS: STIMA-D was designed to provide the profile of patients with major depression in Italy (focusing on pathway to care, patient characteristics, drug therapy and treatment outcomes). The patients enrolled (M/F, aged between 18 and 65) experienced single/multiple episodes of major depression (DSM-IV-TR). Patients with lifetime or current bipolar syndrome or other mental disorders were excluded. RESULTS: 44 of the 50 invited centers sent data concerning 1 140 patients. The majority of patients were women. Among working individuals, 52.5% of them were absent from work due to depression in the previous 6 months. Recurrent episodes of major depression were very common and were associated with persistence of residual post-episodic symptoms, a family history of mood disorders and presence of anxiety. 59.6% of the patients were treated with monotherapy (SSRI or SNRI), while 19.2% of them were treated with SSRI plus SNRI. Only the 25.5% on monotherapy had a complete response compared to 12.4% of patients on dual therapy. DISCUSSION: Poor outcomes in major depression have profound implications on patients' quality of life and cost burden. New pharmacological approaches with novel modes of action are therefore urgently needed.

Cognitive function modulation during aging: a focus on L-alpha-GPE.

OBJECTIVE: The objectives of this review are to explore the neuronal pathways and cellular and molecular mechanisms involved in both healthy and impaired cognitive function and to discuss the role of nootropics, in particular, those with cholinergic activity, as promising interventions to preserve and/or improve cognitive performance in patients in the symptomatic pre-dementia stage, known as mild cognitive impairment (MCI). MATERIALS AND METHODS: Papers were retrieved by a PubMed search, using different combinations of keywords (e.g., cognitive function AND aging AND nootropics), without limitations in terms of publication date or language. RESULTS: Nootropics modulate the activities of specific brain pathways involving neurotransmitters and neuromodulators that have distinct roles in the cognitive processes. The nootropic L-a-glyceryl-phosphoryl-ethanolamine (L-a GPE), by virtue of its action as a phospholipid (PL) precursor and acetylcholine (Ach) donor, targets neural stem cell aging, cholinergic depletion, oxidative stress and microglia activation, loss of entorhinal cortex neurons, and reduced hippocampal volume. Cognitive reserve levels may be linked to the resilience and adaptability of the brain to cope with age-related cognitive decline. L-a GPE may contribute to cognitive reserve preservation via its neuronal well-being promoting action. CONCLUSIONS: The substantial burden of age-related cognitive decline demands effective long-term and well-tolerated interventions aimed at maximizing the span of effective functioning. The use of inappropriate medication may lower cognitive reserve, thus hastening the onset of symptomatic AD, while the use of nootropics, such as L-a GPE may contribute to cognitive reserve preservation via its neuronal well-being promoting action.

Treatment with gut bifidobacteria improves hippocampal plasticity and cognitive behavior in adult healthy rats.

At the present time, gut microbiota inspires great interest in the field of neuroscience as a function of its role in normal physiology and involvement in brain function. This aspect suggests a specific gut-brain pathway, mainly modulated by gut microbiota activity. Among the multiple actions controlled by microbiota at the brain level, neuronal plasticity and cognitive function represent two of the most interesting aspects of this cross-talk communication. We address the possible action of two-months implementation of gut Bifidobacteria using a mixture of three different strains (B-MIX) on hippocampal plasticity and related cognitive behavior in adult healthy Sprague Dawley rats. B-MIX treatment increases the hippocampal BDNF with a parallel gain in dendritic spines' density of hippocampal CA1 pyramidal neurons. Electrophysiological experiments revealed a significant increment of HFS-induced LTP formation on the CA1 hippocampal region in B-MIX treated rats. All these effects are accompanied by a better cognitive performance observed in B-MIX treated animals with no impairments in locomotion activity. Therefore, in adult rats, the treatment with different strains of bifidobacteria is able to markedly enhance neuronal plasticity and the CNS function influencing cognitive behavior, an effect that may suggest a potential therapeutic treatment in brain diseases associated with cognitive functions.

Stimulation of dopamine synthesis in caudate nucleus by intrastriatal enkephalins and antagonism by naloxone.

The intraventricular injection of methionine-enkephalin (50 to 100 micrograms) or [d-Ala2]-methionine-enkephalinamide (1.5 to 12 micrograms), a synthetic enkephalin analog resistant to enzyme degradation, caused a marked dose-dependent increase in dihydroxyphenylacetic acid and homovanillic acid concentrations in the rat striatum. The [d-Ala2] analog increased the accumulation of dopa in the striatum after aromatic amino acid decarboxylase inhibition, indicating that it increased dopamine synthesis. At the highest doses used both enkephalins failed to modify brain serotonin metabolism. The monolateral microinjection of the [d-Ala2]] analog (3 to 6 micrograms) into the caudate nucleus increased the concentration of dihydroxyphenylacetic acid in the injected side, whereas bilateral injection increased the concentration of this compound in both caudate nuclei and caused catalepsy. The stimulant effect of the [d-Ala2] analog on dopamine synthesis in the striatum persisted after destruction of striatal postsynaptic dopamine receptors with kainic acid. The biochemical and behavioral effects of enkephalins were prevented by naloxone, a specific narcotic antagonist. The results indicate that enkephalins stimulate dopamine synthesis by an action on opioid receptors localized on dopaminergic nerve terminals.

Social enrichment reverses the isolation-induced deficits of neuronal plasticity in the hippocampus of male rats.

Environmental enrichment is known to improve brain plasticity and protect synaptic function from negative insults. In the present study we used the exposure to social enrichment to ameliorate the negative effect observed in post weaning isolated male rats in which neurotrophic factors, neurogenesis, neuronal dendritic trees and spines were altered markedly in the hippocampus. After the 4 weeks of post-weaning social isolation followed by 4 weeks of reunion, different neuronal growth markers as well as neuronal morphology were evaluated using different experimental approaches. Social enrichment restored the reduction of BDNF, NGF and Arc gene expression in the whole hippocampus of social isolated rats. This effect was paralleled by an increase in density and morphology of dendritic spines, as well as in neuronal tree arborisation in granule cells of the dentate gyrus. These changes were associated with a marked increase in neuronal proliferation and neurogenesis in the same hippocampal subregion that were reduced by social isolation stress. These results further suggest that the exposure to social enrichment, by abolishing the negative effect of social isolation stress on hippocampal plasticity, may improve neuronal resilience with a beneficial effect on cognitive function.

Isoniazid-induced reduction in GABAergic neurotransmission alters the function of the cerebellar cortical circuit.

The cerebellar cortex contributes to the control of movement, coordination, and certain cognitive functions. The cerebellar network is composed of five different types of neurons that are wired together in a repetitive module. Given that four of these five neurons synthesize and release GABA, this inhibitory neurotransmitter plays a central role in regulation of the excitability and correct functioning of the cerebellar cortex. We have now used isoniazid, an inhibitor of glutamic acid decarboxylase, the enzyme responsible for the synthesis of GABA, to evaluate the contribution of GABAergic transmission in different types of cerebellar cortical neurons to the functioning of the cerebellar circuit. Parasagittal cerebellar slices were prepared from 28- to 40-day-old male rats and were subjected to patch-clamp recording in the voltage- or current-clamp mode. Exposure of the tissue slices to isoniazid (10 mM) resulted in a decrease in the level of GABAergic transmission in Purkinje cells and a consequent increase in the firing rate of spontaneous action potentials that was apparent after 40 min. In granule neurons, isoniazid reduced both tonic and phasic GABAergic currents and thereby altered the flow of information across the cerebellar cortex. Our data support the notion that the amount of GABA at the synaptic level is a major determinant of the excitability of the cerebellar cortex, and they suggest that isoniazid may be a useful tool with which to study the function of the cerebellar network.

Social isolation stress and neuroactive steroids.

Social isolation of rats immediately after weaning is associated to a reduction in the cerebrocortical and plasma concentrations of progesterone and its metabolites 3alpha,5alpha-TH PROG and 3alpha,5alpha-THDOC. Although we found that the basal plasma concentration of adrenocorticotropic hormone in isolated rats was slightly decreased compared with that in group-housed animals no other significant changes were found in the steroidogenic machinery (peripheral benzodiazepine receptors, steroidogenic regulatory protein (StAR)). However, the functional response of the hypothalamic-pituitary-adrenal axis HPA axis to an acute stressful stimulus (foot shock), or to an acute injection of ethanol or isoniazid is markedly increased in isolated rats. Behavioral studies have also indicated that the ability of ethanol to inhibit isoniazid-induced convulsions is greater in isolated rats than in group-housed animals and this effect of isolation is prevented by treatment with the 5alpha-reductase inhibitor finasteride. Social isolation modified the effects of ethanol on the amounts of StAR mRNA and protein in the brain suggesting an alteration in the mechanism of cholesterol transport in mitochondria. Moreover, the amounts of the alpha4 and delta subunits of the GABA(A) receptor in the hippocampus were increased in isolated rats, and these effects were accompanied by an increase in GABA(A) receptor-mediated tonic inhibitory currents in granule cells of the dentate gyrus. Ethanol also increased the amplitude of GABA(A) receptor-mediated miniature inhibitory postsynaptic currents (mIPSC) recorded from CA1 pyramidal neurons with a greater potency in hippocampal slices prepared from socially isolated rats than in those from group-housed, an effect inhibited by finasteride.

GABAergic and dopaminergic transmission in the rat cerebral cortex: effect of stress, anxiolytic and anxiogenic drugs.

Benzodiazepines produce their pharmacological effects by regulating the interaction of GABA with its recognition site on the GABAA receptor complex. In fact, the anxiolytic effect of benzodiazepines may be considered the consequence of the activation of the GABAA receptors induced by these drugs. On the contrary, beta-carboline derivatives which bind with high affinity to benzodiazepine recognition sites modulate the GABAergic transmission in a manner opposite to that of benzodiazepines. Thus, these compounds reduce the function of the GABA-coupled chloride channel and produce pharmacological effects (anxiogenic, proconvulsant and convulsant) opposite to those of benzodiazepines. Taken together, these data strongly indicate that the GABAA receptor complex plays a major role in the pharmacology, neurochemistry and physiopathology of stress and anxiety. This conclusion is further supported by the finding that the function of the GABAA/benzodiazepine receptor complex may be modified by the emotional state of the animals before sacrifice. Accordingly, using an unstressed animal model, the 'handling-habituated' rats, it has been demonstrated that stress, like anxiogenic drugs, decreases the function of GABAA receptor complex, an effect mimicked by the in vivo administration of different inhibitors of GABAergic transmission and antagonized by anxiolytic benzodiazepines. Moreover, a long-lasting down regulation of GABAergic synapses can be obtained after repeated administration of anxiogenic, proconvulsant and convulsant negative modulators of GABAergic transmission. The latter finding further suggests that GABAergic synapses undergo rapid and persistent plastic changes when the GABAergic transmission is persistently inhibited. Finally, the evidence that the activity of mesocortical dopaminergic pathways is altered in opposite manner by drugs that either inhibit or enhance the GABAergic transmission indicates that GABA has a functional role in regulation of dopaminergic neurons in the rat cerebral cortex. Altogether these results suggest that cortical GABAergic and dopaminergic transmission play a major role in the pharmacology, neurochemistry and pathology of the emotional states and fear.

Increase in 20-50 Hz (gamma frequencies) power spectrum and synchronization after chronic vagal nerve stimulation.

OBJECTIVE: Though vagus nerve stimulation (VNS) is an important option in pharmaco-resistant epilepsy, its mechanism of action remains unclear. The observation that VNS desynchronised the EEG activity in animals suggested that this mechanism could be involved in VNS antiepileptic effects in humans. Indeed VNS decreases spiking bursts, whereas its effects on the EEG background remain uncertain. The objective of the present study is to investigate how VNS affects local and inter regional syncronization in different frequencies in pharmaco-resistant partial epilepsy. METHODS: Digital recordings acquired in 11 epileptic subjects 1 year and 1 week before VNS surgery were compared with that obtained 1 month and 1 year after VNS activation. Power spectrum and synchronization were then analyzed and compared with an epileptic group of 10 patients treated with AEDs only. RESULTS: VNS decreases the synchronization of theta frequencies (P < 0.01), whereas it increases gamma power spectrum and synchronization (< 0.001 and 0.01, respectively). CONCLUSIONS: The reduction of theta frequencies and the increase in power spectrum and synchronization of gamma bands can be related to VNS anticonvulsant mechanism. In addition, gamma modulation could also play a seizure-independent role in improving attentional performances. SIGNIFICANCE: These results suggest that some antiepileptic mechanisms affected by VNS can be modulated by or be the reflection of EEG changes.

Morphology, Morphometry and Spatial Distribution of Secondary Osteons in Equine Femur.

A high number of differences exist in bone histological features depending on the species, breed, age and bone. Moreover, osteon distribution may vary in the different sides of a bone as a consequence of different biomechanical strains. The aim of this work was to study the distribution and morphology of osteons in different sides of the equine femoral diaphysis with the attempt to correlate them to the main strains operating on them. The following parameters of secondary osteons and Haversian canals were measured in the transverse sections of diaphyses: perimeter, area, minimum and maximum diameter, eccentricity and osteon population density. A typical Haversian tissue was observed with elliptic secondary osteons consisting in about 10 well-defined lamellae surrounding a circular Haversian canal. Quantitative analysis displays a different population density of secondary osteons depending on the side. The caudal and medial sides, where compression strains are higher, have more secondary osteons in comparison with the cranial and lateral sides, where tension strains are prevalent. These data suggest that secondary osteon population density may depend on the predominant strains. Even the elliptical shape of secondary osteons may be related to biomechanical strains, as their major axes are oriented cranio-caudally parallel to prevalent strains.

D-1 dopamine receptors labelled with 3H-SCH 23390: decrease in the striatum of aged rats.

The effects of age on the binding parameters of 3H-SCH 23390, the most selective D-1 dopamine receptor ligand available at present, were studied in membrane preparations from rat striatum. When compared with 3 month old animals, there was a significant decrease in the density of 3H-SCH 23390 binding sites in 20 month old (-37%) and in 26 month old (-44%) rats, without alterations in the apparent dissociation constant values. No significant changes in the density or affinity of D-1 DA receptors were observed in 14 month old rats. In view of the behavioural effects mediated by D-1 DA receptors recently described (i.e., grooming, stereotypies and EEG desynchronization), the decrease in the density of these receptors in senescent rats may underlie some of the age-related alterations in dopaminergic functions in the rat brain.

Age-related changes of benzodiazepine and GABA binding sites in the rat retina.

The changes in the number and sensitivity of benzodiazepine and GABA binding sites in the rat retina during postnatal development, adulthood and ageing and their functional relationship at different ages have been studied. Data indicate an increase in the total number of both GABA and benzodiazepine binding sites with age. In contrast, the activation of retinal benzodiazepine receptor binding by GABA is significantly reduced in aged rats with respect to young adult and newborn rats. Moreover, the activation of retinal benzodiazepine receptor binding induced by dark exposure of the animals is present in young adult rats but is lost in aged rats. These results suggest that in the retina of aged rats there is an increase of GABA and benzodiazepine receptors which have lost their functional connection.

Functional interaction between benzodiazepine and GABA recognition sites in aged rats.

The present study was undertaken to explore whether there may be age-related changes in benzodiazepine binding and in the functional interaction between GABA and benzodiazepine recognition sites. Data indicate an increase in benzodiazepine binding sites with age. Moreover the functional interactions between GABA and benzodiazepine receptor sites are differentially affected by aging. GABA is less active in enhancing benzodiazepine binding in the older animals because of the loss of GABA receptors, and Diazepam may be more active in enhancing GABA receptor binding in the aged animals because there are more benzodiazepine receptors in this group. An understanding of the relevance of the apparent alteration in coupling between GABA and benzodiazepine receptors must permit a better definition of the behavioral manifestation of their biochemical phenomenon.

Failure of dark adaptation to upregulate D-1 dopamine receptors in retina of senescent rats.

The effect of aging on the binding parameters of 3H-SCH 23390, the most selective ligand of D-1 DA receptors, was studied in membrane preparations from the rat retina. DA-stimulated adenylate cyclase activity was also measured in order to better characterize the changes in retinal D-1 DA receptors induced by aging. The binding studies revealed that the density of 3H-SCH 23390 was increased (34 and 73%) in the retina of 14- and 26-month-old rats, when compared to young adult animals, respectively. In contrast, aging failed to alter the sensitivity of the adenylate cyclase to the action of DA. In fact, DA (10(-6) M to 10(-4) M) elicited a similar enhancement in cyclic AMP formation in retinal homogenates of both adult and senescent rats. Since dark adaptation increases the density of D-1 DA receptors in the retina of adult rats we studied the effect of light deprivation on 3H-SCH 23390 binding and DA-sensitive adenylate cyclase activity in the retina of senescent rats. As previously shown (25) light deprivation increased 3H-SCH 23390 binding and enhanced DA-sensitive adenylate cyclase activity in the retina of young adult rats. On the contrary, dark adaptation failed to increase 3H-SCH 23390 binding and to enhance DA-sensitive adenylate cyclase activity in the retina of senescent rats. Taken together these results indicate that D-1 DA receptors in the retina of aged rats have biochemical and functional properties different from those found in the retina of adult animals; these changes may result in an altered response to the physiological stimuli elicited by environmental lighting.

Involvement of D1 and D2 Dopamine Receptors in the Control of Horizontal Cell Electrical Coupling in the Turtle Retina.

We studied the actions of D1 and D2 dopamine agonists and antagonists on the coupling of horizontal cell axons in the turtle retina by a combination of pharmacological and electrophysiological methods. Both D1 and D2 receptors were identified in membrane fractions by radioligand binding using [3H]-SCH 23390 and [3H]-spiperone, respectively. The KD of both receptor classes were identical (0.21 nM) but D1 receptor density exceeded that of D2 receptors by more than four-fold. D1 agonists increased the activity of adenylate cyclase in a dose-dependent manner, whereas D2 agonists were without significant effect by themselves, nor did D2 antagonists block the D1-mediated increase in adenylate cyclase activity. Intracellular recordings and Lucifer Yellow dye injections were used to characterize the modifications of the receptive field profile of horizontal cell axons (H1AT) exposed to different pharmacological agents. Dopamine or D1 agonists (0.05 - 10 microM) induced a marked constriction of the H1AT receptive field, whereas D2 agonists elicited a small expansion of the receptive field. However, in the presence of a D1 antagonist, as well as IBMX to inhibit phosphodiesterase, D2 agonists (10 - 70 microM) induced a marked increase in the receptive field profile. These results indicate that both D1 and D2 dopamine receptors play a role in shaping the receptive field profile of the horizontal cell axon terminal in the turtle retina.

Propofol in anesthesia. Mechanism of action, structure-activity relationships, and drug delivery.

Propofol (2,6-diisopropylphenol) is becoming the intravenous anesthetic of choice for ambulatory surgery in outpatients. It is extensively metabolized, with most of the administered dose appearing in the urine as glucuronide conjugates. Favorable operating conditions and rapid recovery are claimed as the main advantages in using propofol, whereas disadvantages include a relatively high incidence of apnea, and blood pressure reductions. Besides a literature summary of the pharmacodynamics, pharmacokinetics, toxicology, and clinical use, this review provides a deeper discussion on the current understanding of mechanism of action and structure-activity relationships, and recent findings on drug delivery technologies as applied to the improvement of propofol formulations. The action of propofol involves a positive modulation of the inhibitory function of the neurotransmitter gama-aminobutyric acid (GABA) through GABAA receptors. Recent results from recombinant human GABAA receptor experiments and findings from the work exploring the effects at other receptors (e.g., glycine, nicotinic, and M1 muscarinic receptors) are reviewed. Studies showing its antiepileptic and anxiolytic properties are also discussed. The structure-activity relationships (SAR) of series of alkylphenols and p-X-substituted congeners have been reinvestigated. Interestingly, unlike the other congeners tested sofar, p-iodo-2,6-diisopropylphenol displayed anticonvulsant and anticonflict effects, but not sedative-hypnotic and anesthetic properties. Due to its high lipid-solubility, propofol was initially formulated as a solution with the surfactant Cremophor EL, but the occurrence of pain on injection and anaphylactoid reactions prompted to search for alternative formulations. Results from using cyclodextrins, water-soluble prodrugs, and adopting Bodor's approach to the site-specific chemical delivery system (CDS), as well as the advantages provided by computer-controlled infusion systems, are examined in some detail.

Clozapine, but not haloperidol, increases brain concentrations of neuroactive steroids in the rat.

The extrapyramidal side effects of typical antipsychotics, which are induced to a markedly reduced extent by clozapine, have been linked to a dysfunction of central gamma-aminobutyric acid (GABA)-mediated neurotransmission. The effects of clozapine on the brain concentrations of 3alpha-hydroxy-5alpha-pregnan-20-one (allopregnanolone, AP) and 3alpha,21-dihydroxy-5alpha-pregnan-20-one (allotetrahydrodeoxycorticosterone, THDOC), two potent and endogenous positive allosteric modulators of GABA-mediated chloride current intensities at GABA(A) receptors, were compared with those of the typical antipsychotic haloperidol. A single administration of clozapine (1.25-20 mg/kg, IP), but not of haloperidol (0.1 or 0.5 mg/kg, IP), induced dose- and time-dependent increases in the concentrations of progesterone, AP, and THDOC in the cerebral cortex and striatum of rats. Clozapine (at 10 mg/kg, but not at lower doses) also increased the concentrations of these steroids as well as that of corticosterone in plasma in intact rats, but failed to increase the cortical concentrations of AP and THDOC in adrenalectomized-orchidectomized rats. An acute challenge with clozapine (10 mg/kg), administered 48 h after the termination of daily treatment with the same dose for 19 days, still increased the cortical concentrations of progesterone, AP, and THDOC. These results suggest that the clozapine-induced increases in neuroactive steroid concentrations in the brain may contribute to the atypical pharmacological profile of this antipsychotic drug.

Proconflict effect of GABA receptor complex antagonists. Reversal by diazepam.

The effect of drugs which down-regulate the function of GABA at the level of the GABA/benzodiazepine receptor complex was studied on the conflict test in the rat. The GABA receptor antagonist, bicuculline, and the blockers of the GABA-receptor-coupled chloride channel, picrotoxin and pentylenetetrazol, produced a dose-dependent proconflict effect. This effect occurred at dose levels which failed to affect unpunished behaviour. The most effective compounds were bicuculline and picrotoxin. The proconflict effect of these drugs was prevented by diazepam but not by the specific benzodiazepine antagonist, Ro15-1788. The data indicate that a diminished GABAergic activity at different subunits of the GABA receptor complex resulted in an enhancement of punishment-suppressed behaviour in rats.

Changes in GABA(A) receptor gene expression induced by withdrawal of, but not by long-term exposure to, zaleplon or zolpidem.

The effects of long-term treatment with and subsequent withdrawal of the two hypnotic drugs zaleplon and zolpidem on the abundance of gamma-aminobutyric acid type A (GABA(A)) receptor subunit mRNAs in cultured rat cerebellar granule cells were investigated. Incubation of neurons with either drug at a concentration of 10 microM for 5 days did not significantly affect the amounts of mRNAs encoding the alpha(1), alpha(4), beta(1), beta(2), beta(3), gamma(2)L, or gamma(2)S subunits. As expected, similar treatment with the nonselective benzodiazepine diazepam resulted in a decrease in the abundance of alpha(1), beta(2), gamma(2)L, and gamma(2)S subunit mRNAs as well as an increase in that of the beta(1) subunit mRNA. Withdrawal of zaleplon or zolpidem, like that of diazepam, induced a marked increase in the amount of the alpha(4) subunit mRNA. In addition, discontinuation of treatment with either hypnotic drug resulted in a decrease in the amounts of alpha(1), beta(2), gamma(2)L, and gamma(2)S subunit mRNAs as well as an increase in that of the beta(1) subunit mRNA. These effects of zaleplon and zolpidem on GABA(A) receptor gene expression are consistent with the reduced tolerance liability of these drugs, compared with that of diazepam, as well as with their ability to induce both physical dependence and withdrawal syndrome.

Binding sites for [3H]2-oxo-quazepam in the brain of the cat: evidence for heterogeneity of benzodiazepine recognition sites.

In the present study, the distribution of benzodiazepine recognition site subtypes in the brain of the cat was investigated. To this aim, the binding properties of [3H]2-oxo-quazepam ([3H]2OXOQ) and [3H]beta-CCE, two ligands with preferential affinity for Type I benzodiazepine recognition sites, were compared to binding parameters for [3H]flunitrazepam ([3H]FNT) in different areas of the cat brain. The ratio of [3H]2OXOQ to [3H]FNT binding sites indicated that, in the cerebellum, Type I sites accounted for 90% of the total number of benzodiazepine recognition sites. The cerebral cortex, thalamus and mesencephalic reticular formation had also a high proportion of Type I sites (73-78%), whilst the two subtypes were almost equally distributed in the hippocampus, amygdala and bulbar reticular formation. A similar distribution of subtypes of benzodiazepine recognition sites was indicated by the ratio of [3H]beta CCE to [3H]FNT binding sites for different areas of the brain. These results demonstrate the existence of heterogeneity of recognition sites for benzodiazepines in the brain of the cat and support the view that [3H]2OXOQ preferentially labels Type I sites.