Elena Romeo

Plasma Concentrations of Neuroactive Steroids before and after Repetitive Transcranial Magnetic Stimulation (rTMS) in Major Depression

There is evidence for altered levels of neuroactive steroids in major depression that normalize after successful antidepressant pharmacotherapy. Currently it is not known whether this is a general principle of clinically effective antidepressant therapy or a pharmacological effect of antidepressants. Here, we investigated whether repetitive transcranial magnetic stimulation (rTMS) may affect plasma concentrations of neuroactive steroids in a similar way as antidepressant pharmacotherapy. Progesterone, 3α,5α-tetrahydroprogesterone (3α,5α-THP), 3α,5β- tetrahydroprogesterone (3α,5β–THP), 3β,5α-tetrahydroprogesterone (3β, 5α-THP) and dehydroepiandrosterone (DHEA) were quantified in 37 medication-free patients suffering from a major depressive episode before and after 10 sessions of left prefrontal rTMS. Plasma samples were analyzed by means of a highly sensitive and specific combined gas chromatography/mass spectrometry analysis. There was a significant reduction of depressive symptoms after rTMS. However, plasma concentrations of neuroactive steroids were not affected by rTMS and not related to clinical response. Clinical improvement after extended daily treatment with rTMS is not accompanied by changes in neuroactive steroid levels. Changes in neuroactive steroid levels after antidepressant pharmacotherapy more likely reflect specific pharmacological effects of antidepressant drugs and are not necessary for the amelioration of depressive symptoms.

Stimulation of Brain Pregnenolone Synthesis by Mitochondrial Diazepam Binding Inhibitor Receptor Ligands In Vivo

Abstract: Evidence that neurosteroids are potent modulators of the action of GABA at GABAA receptors has prompted the investigation of the mechanism that controls brain neurosteroid synthesis by glial cell mitochondria in vivo. In vitro studies suggest that the interaction of the diazepam binding inhibitor (DBI)—a polypeptide that is abundant in steroidogenic cells—with glial mitochondrial DBI receptors (MDRs) is a crucial step in the physiological regulation of neurosteroid biosynthesis. MDRs bind 4‐chlorodiazepam (4′‐CD), N,N‐di‐n‐hexyl‐2‐(4‐fluorophenyl)‐indol‐3‐acetamide (FGIN‐1–27), and the isoquinoline carboxamide PK 11195 with high affinity, and these ligands have been used to investigate whether the stimulation of glial MDRs increases brain pregnenolone production in vivo. Adrenalectomized and castrated (A‐C) male rats (to eliminate peripheral sources of pregnenolone) were pretreated with trilostane (to prevent pregnenolone metabolism to progesterone), and the pregnenolone content in brain regions dissected after fixation with a 0.8‐s exposure to microwave irradiation focused to the head was determined by HPLC followed by specific radioimmunoassay. The forebrain and cerebellum of A‐C rats contained 4–7 ng of pregnenolone/g of tissue, and the olfactory bulb contained 10–14 ng/g. These concentrations of brain pregnenolone are only 30–40% lower than those of shamoperated rats. In contrast, the plasma pregnenolone content of sham‐operated rats was 2–3 ng/ml, but it was only 0.15–0.20 ng/ml in the plasma of A‐C rats. In A‐C rats, treatment with the MDR ligands 4‐CD and FGIN‐1–27 increased the pregnenolone content in the brain but failed to change the plasma or peripheral tissue content of this steroid. The effect of 4′‐CD on brain pregnenolone content was maximal (70–100% increase) at the dose of 18 μmol/kg, 5–10 min after intravenous injection. The effect of oral administration of FGIN‐1–27 on brain pregnenolone content was maximal (80–150% increase) at doses of 400–800 μmollkg and peaked at ∼ 1 h. That this effect of FGIN‐1–27 was mediated by the MDR was documented by pre‐treatment with the MDR partial agonist PK 11195 (100 μmol/kg, i.p.). PK 11195 did not affect basal brain pregnenolone content but prevented the accumulation of brain pregnenolone induced by FGIN‐1–27. FGIN‐1–27 and 4‐CD failed to increase the brain concentration of dehydre epiandrosterone in A‐C rats. These data suggest that glial cell MDRs play a role in neurosteroid biosynthesis in vivo.

The pharmacology of neurosteroidogenesis

In adrenal cortex and other steroidogenic tissues including glial cells, the conversion of cholesterol into pregnenolone is catalyzed by the cytochrome P450scc located in the inner mitochondrial membrane. A complex mechanism operative in regulating cholesterol access to P450scc limits the rate of pregnenolone biosynthesis. Participating in this mechanism are DBI (diazepam binding inhibitor), an endogenous peptide that is highly expressed in steroidogenic cells and some of the DBI processing products including DBI 17-50 (TTN). DBI and TTN activate steroidogenesis by binding to a specific receptor located in the outer mitochondrial membrane, termed mitochondrial DBI receptor complex (MDRC). MDRC is a hetero-oligomeric protein: only the subunit that includes the DBI and benzodiazepine (BZD) recognition sites has been cloned. Several 2-aryl-3-indoleacetamide derivatives (FGIN-1-X) with highly selective affinity (nM) for MDRC were synthesized which can stimulate steroidogenesis in mitochondrial preparations. These compounds stimulate adrenal cortex steroidogenesis in hypophysectomized rats but not in intact animals. Moreover, this steroidogenesis is inhibited by the isoquinoline carboxamide derivative PK 11195, a specific high affinity ligand for MDRC with a low intrinsic steroidogenic activity. Some of the FGIN-1-X derivatives stimulate brain pregnenolone accumulation in adrenalectomized-castrated rats. The FGIN-1-X derivatives that increase brain pregnenolone content, elicit antineophobic activity and antagonize punished behavior in the Vogel conflict test in rats. These actions of FGIN-1-X are resistant to inhibition by flumazenil, a specific inhibitor of BZD action in GABAA receptors but are antagonized by PK 11195, a specific blocker of the steroidogenesis activation via MDRC stimulation. It is postulated that the pharmacological action of FGIN-1-X depends on a positive modulation of the GABA action on GABAA receptors mediated by the stimulation of brain neurosteroid production.

The complex roles of neurosteroids in depression and anxiety disorders.

The role of neurosteroids in neuropsychiatric disorders has been thoroughly investigated in many research studies that have stressed their significant pathophysiological function in neuropsychiatry. In this review, we will focus mainly on the steroids active on the GABA(A) receptors studied in anxiety and depression. The aim is to discuss the controversial results reported in research on anxiety and depressive disorders. We suggest the combined use of biological parameters linked to psychopathological dimensions to make more homogeneous diagnoses and to develop more precise therapies for the treatment of depression and anxiety disorders. We discuss the role of neurosteroids in the pathophysiology and therapy of anxiety and depression. Finally, we consider the possibility of using quantification of mRNA expression of steroidogenic enzymes from peripheral sources in neuropsychiatry.

Panic Induction with Cholecystokinin-Tetrapeptide (CCK-4) Increases Plasma Concentrations of the Neuroactive Steroid 3|[alpha]|, 5|[alpha]| Tetrahydrodeoxycorticosterone (3|[alpha]|, 5|[alpha]|-THDOC) in Healthy Volunteers

3α-reduced neuroactive steroids such as 3α, 5α-tetrahydroprogesterone (3α, 5α-THP) and 3α, 5α-tetrahydrodeoxycorticosterone (3α, 5α-THDOC) are potent positive allosteric modulators of γ-aminobutyric acid type A (GABAA) receptors and display pronounced anxiolytic activity in animal models. Experimental panic induction with cholecystokinin-tetrapeptide (CCK-4) and sodium lactate is accompanied by a decrease in 3α, 5α-THP concentrations in patients with panic disorder, but not in healthy controls. However, no data are available on 3α, 5α-THDOC concentrations during experimental panic induction. Therefore, we quantified 3α, 5α-THDOC concentrations in 10 healthy volunteers (nine men, one woman) before and after panic induction with CCK-4 by means of a highly sensitive and specific gas chromatography/mass spectrometry analysis. CCK-4 elicited a strong panic response as assessed by the Acute Panic Inventory. This was accompanied by an increase in 3α, 5α-THDOC, ACTH and cortisol concentrations. This increase in 3α, 5α-THDOC might be a consequence of hypothalamic–pituitary–adrenal (HPA) axis activation following CCK-4-induced panic, and might contribute to the termination of the anxiety/stress response following challenge with CCK-4 through enhancement of GABAA receptor function.

Plasma concentrations of neuroactive steroids before and after electroconvulsive therapy in major depression

There is evidence that both cerebrospinal fluid (CSF) and plasma concentrations of 3α-reduced neuroactive steroids are decreased in major depressive disorder. Successful antidepressant pharmacotherapy, for example, with selective serotonin reuptake inhibitors (SSRIs), over several weeks is accompanied by an increase in CSF and plasma concentrations of these neuroactive steroids. However, no such increase has been observed during nonpharmacological treatments such as partial sleep deprivation or repetitive transcranial magnetic stimulation. In order to investigate whether concentration changes in neuroactive steroids are an important component of clinically effective antidepressant treatment, we examined plasma concentrations of the neuroactive steroids 3α,5α-tetrahydroprogesterone, 3α,5β-tetrahydroprogesterone, 3β,5α-tetrahydroprogesterone, and their precursors progesterone, 5α-dihydroprogesterone, and 5β-dihydroprogesterone in 31 pharmacotherapy-resistant depressed in-patients before and after unilateral electroconvulsive therapy (ECT) as a monotherapy over 4 weeks. Samples were quantified for neuroactive steroids by means of a highly sensitive and specific combined gas chromatography/mass spectrometry analysis. In all, 51.6% of the patients were treatment responders. There was no influence of ECT on the plasma concentrations of any neuroactive steroid studied. Moreover, neuroactive steroid levels did not differ between treatment responders and nonresponders. Our study shows that changes in neuroactive steroid plasma levels are not a mandatory factor for successful antidepressant treatment by ECT. Thus, the previously observed changes in plasma concentrations of neuroactive steroids following treatment with antidepressants such as SSRIs more likely reflect distinct pharmacological properties of these compounds rather than clinical improvement.

Neuroactive Steroids in Depression and Anxiety Disorders: Clinical Studies

Certain neuroactive steroids modulate ligand-gated ion channels via non-genomic mechanisms. Especially 3α-reduced pregnane steroids are potent positive allosteric modulators of the γ-aminobutyric acid type A (GABAA) receptor. During major depression, there is a disequilibrium of 3α-reduced neuroactive steroids, which is corrected by clinically effective pharmacological treatment. To investigate whether these alterations are a general principle of successful antidepressant treatment, we studied the impact of nonpharmacological treatment options on neuroactive steroid concentrations during major depression. Neither partial sleep deprivation, transcranial magnetic stimulation, nor electroconvulsive therapy affected neuroactive steroid levels irrespectively of the response to these treatments. These studies suggest that the changes in neuroactive steroid concentrations observed after antidepressant pharmacotherapy more likely reflect distinct pharmacological properties of antidepressants rather than the clinical response. In patients with panic disorder, changes in neuroactive steroid composition have been observed opposite to those seen in depression. However, during experimentally induced panic induction either with cholecystokinine-tetrapeptide or sodium lactate, there was a pronounced decline in the concentrations of 3α-reduced neuroactive steroids in patients with panic disorder, which might result in a decreased GABAergic tone. In contrast, no changes in neuroactive steroid concentrations could be observed in healthy controls with the exception of 3α,5α-tetrahydrodeoxycorticosterone. The modulation of GABAA receptors by neuroactive steroids might contribute to the pathophysiology of depression and anxiety disorders and might offer new targets for the development of novel anxiolytic compounds.

Neurosteroid and neurotransmitter alterations in Parkinson's disease

Parkinsons disease (PD) is associated with a massive loss of dopaminergic cells in the substantia nigra leading to dopamine hypofunction and alteration of the basal ganglia circuitry. These neurons, are under the control, among others, of the excitatory glutamatergic and inhibitory γ-aminobutyric acid (GABA) systems. An imbalance between these systems may contribute to excitotoxicity and dopaminergic cell death. Neurosteroids, a group of steroid hormones synthesized in the brain, modulate the function of several neurotransmitter systems. The substantia nigra of the human brain expresses high concentrations of allopregnanolone (3α, 5αtetrahydroprogesterone), a neurosteroid that positively modulates the action of GABA at GABAA receptors and of 5α-dihydroprogesterone, a neurosteroid acting at the genomic level. This article reviews the roles of NS acting as neuroprotectants and as GABAA receptor agonists in the physiology and pathophysiology of the basal ganglia, their impact on dopaminergic cell activity and survival, and potential therapeutic application in PD.

Increased behavioral neurosteroid sensitivity in a rat line selectively bred for high alcohol sensitivity

Acute administration of a neurosteroid 5beta-pregnan-3alpha-ol-20-one induced a greater impairment in motor performance of the selectively bred alcohol-sensitive (ANT) than alcohol-insensitive (AT) rats. This difference was not associated with the sensitivity of gamma-aminobutyrate type A (GABA(A)) receptors, as 5alpha-pregnan-3alpha-ol-20-one (allopregnanolone) decreased the autoradiographic signals of t-butylbicyclophosphoro[35S]thionate binding to GABA(A) receptor-associated ionophores more in the brain sections of AT than ANT rats. Nor was the difference associated with baseline levels of neuroactive progesterone metabolites, as 5alpha-pregnan-3,20-dione (5alpha-DHP) and 5alpha-pregnan-3alpha-ol-20-one were lower in the ANT rats. After ethanol (2 g/kg, i.p.) administration and the subsequent motor performance test, the increased brain concentrations of these metabolites were still lower in the ANT than AT rats, although especially in the cerebellum the relative increases were greater in the ANT than AT rats. The present data suggest that the mechanisms mediating neurosteroid-induced motor impairment are susceptible to genetic variation in rat lines selected for differences in ethanol intoxication.

Changes in CCK-4 induced panic after treatment with the GABA-reuptake inhibitor tiagabine are associated with an increase in 3α,5α-tetrahydrodeoxycorticosterone concentrations

There is evidence that gamma-amino-butyric acid type A (GABA(A))-receptor modulating neuroactive steroids play a role in the pathophysiology of panic disorder. Antidepressant treatment has been suggested to stabilize the concentrations of neuroactive steroids. In this pilot study we investigated neuroactive steroid concentrations during GABAergic treatment, which might represent an alternative anxiolytic pharmacotherapeutic strategy. Neuroactive steroid concentrations were determined in 10 healthy subjects treated with tiagabine. To evaluate the anxiolytic effects of tiagabine a cholecystokinin-tetrapeptide (CCK-4) challenge was performed before and after treatment. Treatment with tiagabine led to a significant increase in 3alpha,5alpha-tetrahydrodeoxycorticosterone (3alpha,5alpha-THDOC) from 0.49 to 1.42 nmol/l (Z=-2.80, p=.005), which was significantly correlated with a decrease of panic symptoms in the CCK-4 challenge. Thus, it might be hypothesized that the anxiolytic effects of GABAergic treatment might in part be mediated by their influence on 3alpha,5alpha-THDOC concentrations.