Neuroactive steroids

AHopregnanolone and similar A-ring-reduced pregnanes potentiate GABA effects at these receptors. These steroids mimic the effects of the benzodiazepines, changing chloride ion conductance and producing sedative and hypnotic behavioral effects (276,277). Neuroactive steroids can be therapeutically useful as anticonvulsants, anxiolytics, or anesthetics (qv) (see also Hypnotics, sedatives, anticonvulsants, and anxiolytics). 

Glycine receptor function is modulated by alcohols and anesthetics [4]. Amino acid residue al(S267) is critical for alcohol potentiation, as mutation to small residues (Gly, Ala) enhance, and mutation to large residues (His, Cys, Tyr) diminish the ethanol effect. Glycine recqrtor modulation by Zn2+ involves structural determinants located within the large N-terminal domain. Additional glycinergic modulators include neuroactive steroids and the anthelmintic, ivermectin, which activates glycine receptors by a novel, strychnine-insensitive mechanism. 

Neurosteroids are neuroactive steroids, which are synthesized in the brain. Neurosteroids can bind to and modulate the activity of y-aminobuty tie acidA(GABA A) receptors. 

With so many different neurosteroids with differing and even opposing neuronal effects, much will depend on their relative concentrations at any time and any evaluation of their function must take this into consideration. Hopefully the synthesis and use of appropriate antagonists will throw more light on the physiological role of steroids in the CNS and facilitate the development and clinical use of new neuroactive steroids (see Gasior et al. 1999). 

Gasior, M, Carter, RB and Witkin, JM (1999) Neuroactive steroids potential therapeutic use in neurological and psychiatric disorders. Trends Pharmacol. Sci. 20 107-112. 

Rupprecht R and Holsboer F (1999) Neuroactive steroids mechanisms of action and neuropsychopharmacological perspectives. Trends Neurosci. 22 410-416. 

Porcu, P., Sogliano, C., Cinus, M., Purdy, R.H., Biggio, G., Concas, A. Nicotine induced changes in cerebrocortical neuroactive steroids and plasma corticosterone concentrations in the rat. Pharmacol. Biochem. Behav. 74 683, 2003. 

While steroids generated in the brain have been referred to as neurosteroids , another useful term is neuroactive steroids to refer to all steroids that affect brain function via any mechanism and irrespective of site of formation. The term neuroactive steroid also has been used to describe neuroactive steroid drugs. 

Estradiol. The first neuroactive steroid receptor type to be recognized was that for estradiol [3]. In vivo uptake of [3H] estradiol, and binding to cell nuclei isolated from hypothalamus, pituitary and other brain regions, revealed steroid specificity closely resembling that of the uterus, where steroid receptors were first discovered [3]. Cytosolic estrogen receptors isolated from pituitary and brain tissue closely resemble those found in uterus and mammary tissue. A hallmark of the estrogen receptor is its existence... 

Mihalek, R. M., Banerjee, P. K., Korpi, E. R., and Quinlan, J. J. (1999) Attenuated sensitivity to neuroactive steroids in y-aminobutyrate type A receptor 8 subunit knockout mice. Proc. Natl. Acad. Sci. USA 96,12905-12910. 

Nguyen TT, Matsumoto K, Yamasaki K, Watanabe H. (1997). Majonoside-R2 reverses social isolation stress-induced decrease in pentobarbital sleep in mice possible involvement of neuroactive steroids. Life Sciences. 61(4) 395-402. 

Purdy RH, Fitzgerald RL, Everhart ET, Mellon SH, Alomary A, Parsons LH. 2004. The analysis of neuroactive steroids by mass spectrometry. Handbook of Neurochemistry and Molecular Neurobiology, 3rd edition. Volume 18 Practical Neurochemistry (Methods). Baker GB, Dunn SMJ, Holt A, editors. New York Kluwer Academic Publishers. 

A prevailing problem with neuroactive steroid analysis by HPLC is the lack of sufficient chromaphores or fluorophores within their chemical structures to allow suitable spectrophotometric end-point detection such as with UV/VIS or fluorescence with adequate sensitivity. The multitude of structural isomers of the metabolites also decreases the applicability of RP-HPLC since the chromatographic profiles become very complex with co-eluting peaks. Due to these inherent problems, it is often necessary to derivatize this group of compounds prior to chromatographic separation and suitable end-point detection to allow their direct determination at physiological concentrations. 

A variety of endogenous and synthetic steroids have now been demonstrated to alter neuronal activity. Paul and Purdy (1992) proposed that this group of neurosteroids be named neuroactive steroids, a nomenclature defining their activity. Further complicating this terminology, some authors more generally define neurosteroids both as neuroactive compounds produced de novo in the nervous system and as those steroids derived from circulating precursors, which are metabolized to neuroactive compounds in the nervous system. 

In our laboratories, we have abandoned using RIA for the measurement of neuroactive steroids except where the RIA procedure has been strictly validated by prior MS identification for each experimental protocol. Immunoassays that measure low concentrations of steroids in complex biological matrices are... 

Neuroactive steroids are comparatively polar compounds that, in most cases, are derivatized by a suitable reagent before being analyzed by GC/MS. It is essential to add functional groups that enhance ionization efficiency, decrease polarity, and increase volatility of the steroid, thus making it more easily detected by GC/MS. Suitable derivatization of neuroactive steroids also results in enhanced sensitivity and specificity of mass spectra. 

Selected ion monitoring GC/MS trace of the HFBA derivatives of nine neuroactive steroids found in human plasma. The separations were accomplished on a Restek Rtx-200 MS column. Relative absorbance was measured at negative Cl for m/z values of 474 (pregnanolones), 492 (pregnenolone), 664 (androstanediols), and 706 (pregnanediolones). (Everhart et al., unpublished)... 

The recent development of mixed-mode stationary phases may provide new approaches for the separation of neuroactive steroids from biological matrices. These stationary phases typically embed a charged functional group within the nonpolar carbon chains typically employed in RP separations, and as such separations based on both RP and ion-exchange characteristics can be designed. Examples of this column type are the Primesep line of columns from SIELC Technologies. This separation approach maybe quite useful for the separation of steroid sulfates from complex mixtures. 

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