neurosteroids ( ns ) , a term proposed by the physiologists baulieu and robel ( 1990 ) , is widely used to refer to the steroids synthesized in the brain . through their interaction with neuronal membrane receptors and ion channels ,
they are capable to modify the brain excitability ( lambert et al . , 2003 ;
depending on its chemical structure , the steroids interactions with the gabaa receptor may produce positive or negative modulations ( majewska , 1992 ; reddy , 2003 ) . among the positive modulators of this receptor are two progesterone 's metabolites : the 5-pregnane-3-ol-20-one ( allopregnanolone ) and its isomer 5-pregnane-3-ol-20-one ( pregnanolone ; gasior et al . , 1999 ) .
the interest on these steroids arises from their potential activity as anticonvulsants , anesthetics , anxiolytic or sedative - hypnotic agents ( akk et al . , 2007 ) useful for the treatment of several neurological and psychiatric disorders ( gasior et al . , 1999 ) .
also , various physiological and pathophysiological conditions have been associated with changes in allopregnanolone and pregnanolone levels ( akk et al . , 2007 ) .
although the natural ns can be used in epileptic patients ( herzog , 1999 ) , certain properties , like their short biological half - life , avoid their clinical use .
for that reason , synthetic steroids ( ss ) , that exhibit better bioavailability and efficacy , have an important therapeutic potential in brain disorders , becoming an alternative for this kind of pathologies ( reddy and kulkarni , 2000 ; morrow , 2007 ) .
the medicinal chemistry of neuroactive steroids ( nas ) has been focused in the development of ss analogues preserving the absolute configuration of naturally occurring steroids .
structure / activity studies indicate that the 3-hydroxyl configuration is required for binding and activity ( purdy et al . ,
for example , the 3-methylated synthetic analog of allopregnanolone , ganaxolone ( 3-hydroxy-3-methyl-5-prengan-20-one ) is capable to overcome these limitations , showing effective anticonvulsant properties ( carter et al . , 1997 ; reddy and woodward , 2004 ) .
in fact , until now , it is the only ss that has been proved in human clinical trials for epilepsy ( nohria et al . , 2010 ) .
according to the reversal potential of the permeate ions , the postsynaptic gaba response can be excitatory or inhibitory ( akk et al . , 2007 ) .
the binding of the convulsant t - butyl - bicyclophosphorothionate ( tbps ) to the gabaa receptor can be allosterically modulated by allopregnanolone and pregnanolone ( ramanjaneyulu and ticku , 1984 ) . when gaba is present , these metabolites have a significantly increased binding affinity , and under this condition , it is possible to reflect the functional state of this receptor ( majewska , 1992 ; hawkinson et al . , 1994 ) .
similarly , nas can also stimulate the binding of flunitrazepam or muscimol to the receptor ( majewska et al .
the ns exposure enhances the opening probability of the chloride channel , so that the mean time open is increased , resulting in a reduction of neuronal excitability .
harrison and simmons ( 1984 ) demonstrated that alphaxalone ( alpx ; 3-hydroxy-5-pregnane-11,20-dione ) , another allopregnanolone synthetic analogue , was able to enhance the gaba - evoked responses .
also , a positive allosteric modulation of gabaa receptor was found with the ss ganaxolone ( carter et al . , 1997 ; gasior et al . , 1997
it has been described that at least two ent-16-ketosteroid synthetic analogues ( 3-5-androsten-16-one and 3-5-4methoxy - androsten-16-one ; with an absolute opposite configuration to nas ) , produced a more potent inhibition of the tbps binding than alpx ( qian et al . , 2013 ) .
moreover , we showed a decrease in tbps binding and an increase in flunitrazepam and muscimol binding by the administration of ss epoxies ( analogues to allopregnanolone and pregnanolone ) with an intramolecular oxygen bridge that keeps the a / b angle of the steroid nucleus in a controlled way ( veleiro and burton , 2009 ; rey et al . , 2013 ) .
cumulative evidence indicates the existence of neuroprotective properties of nas in a variety of experimental paradigms ( schumacher et al . , 2004 ) .
they have a major influence on the central nervous system ( cns ) activity and are essential for growth and survival of neurons and glial cells ( wang et al . , 2005 ; melcangi et al . ,
studies in adult animals after brain injury indicate that nas have an important role in repairing processes , enhancing myelination and reducing apoptotic processes ( ibanez et al . , 2004 ) .
during pregnancy , stressful events which lead to transient hypoxia / ischemia , stimulate nas production in the brain providing further protection ( nguyen et al . , 2004 ) .
this supports the importance of nas in brain development and suggests that the exposure to normal nas levels is critical . in traumatic brain injury
, progesterone has the most important repair - promoting actions ( he et al . , 2004a ) and
it acts through its reduced metabolites like allopregnanolone ( djebaili et al . , 2004 ; he et al .
the neuroprotective actions of allopregnanolone have been shown in hypoxia - induced brain injury models , where its levels increase in response to acute hypoxic stress , as a protective mechanism to reduce excitotoxicity ( hirst et al . , 2006 ) .
in fact , we have described a protective effect of allopregnanolone on astrogliosis ( kruse et al . , 2009 ) and
neuronal damage ( kruse et al . , 2010 ) caused by hypoxia in perinatal cultures of cerebral cortex and hippocampus of the rat .
studies with the ss mifepristone ( ru486 ) , reported that it acts as a neuroprotective agent against excitotoxicity and traumatic brain injury ( behl et al .
, 1997 ; mccullers et al . , 2002 ) and protects purkinje cells from cell death in postnatal rat and mouse cerebellum organotypic slice cultures ( ghoumari et al . , 2003 ) , through the reversion of chloride efflux in the gabaa receptor elicited by gaba ( rakotomamonjy et al . ,
we have also demonstrated that two ss epoxies , ( analogues of allopregnanolone and pregnanolone , ) were capable to prevent the glial and neuronal damages in the perinatal cultures of cerebral cortex and hippocampus ( rey et al . , 2013 ) . in adults , the brain ischemic stroke
the loss of energy supply by the mitochondrial dysfunction and posterior increased oxidative stress contributed to the neuronal injury .
therefore , a trend has been set in the development of steroid drugs that reduce the excitotoxicity and the oxidative stress , for treatments of acute brain injuries or chronic neurodegenerative diseases . because the current therapies are still limited
one example is the ss 5-androst-3,5,6-triol showed a robust neuroprotective effects when it was tested in vitro ( chen et al . , 2013 ) .
the alzheimer 's disease ( ad ) produces a brain degenerative process , with neuronal losses and decreased synapses .
present therapies are focused on stopping the progression of the disease , but the major challenge remains , in restore cognitive function through the regeneration of lost neurons and neural circuitry . in aged and ad brains , the pool of neural stem cells , their proliferative potential and the allopregnanolone content are markedly diminished ( bernardi et al . , 1998 ; genazzani et al . , 1998 ; weill - engerer et al . ,
studies using transgenic ad mice showed that allopregnanolone has neurogenic properties ( wang et al . , 2008 ) .
these in vitro and in vivo neurogenic features , coupled to low molecular weight , easy blood brain barrier penetration and lack of toxicity , are the key elements required to consider the use of allopregnanolone as a neurogenic / regenerative therapy for neurons restoration in ad patients ( brinton and wang , 2006 ; irwin and brinton , 2014 ) .
estrogen has also showed neuroprotective properties , preventing the development of neurodegenerative disorders like ad .
hormonal therapy at menopause ( to restore normal levels ) appears to reduce the risks , but this kind of treatment has been associated with detrimental effects .
moreover , estrogen like neuroprotection effects were observed with the ss 4-estren-3,17-diol that differs structurally from estrogens only on the a ring ( kousteni et al . , 2002 ; cordey et al . ,
in addition , similar neuroprotective actions have been described with the ss ent - steroid of 17-estradiol ( covey , 2009 ) .
another important issue is the influence of the ss on the local natural ns synthesis .
nas are present in the nervous system and in other steroidogenic tissues , like gonads and adrenal glands . in the cns ,
, the cholesterol is converted to pregnenolone by the cytochrome p450 side - chain cleavage enzyme ( cyp450scc ; iwahashi et al . ,
then , the pregnenolone is oxidized to progesterone by the 3-hydroxysteroid dehydrogenase enzyme ( 3-hsd ; zwain and yen , 1999 ) being this conversion an essential step in the biosynthesis of all steroid hormones .
allopregnanolone is synthesized from progesterone , by the sequential enzymatic steps of the type i 5-reductase ( 5-r ) and the 3-hydroxysteroid dehydrogenase enzymes ( 3-hsd ; mellon et al . , 2001 ) .
the rate - limiting step in neurosteroidogenesis is the unidirectional reduction of progesterone to the 5-dihydroprogesterone ( 5-dhp ) by the 5-r .
functionally expression of these enzymes has been described in pluripotent progenitor cells ( melcangi et al . ,
the expression of 3-hsd enzyme has been demonstrated in several tissues like adrenal glands , gonads and cns ( rheaume et al . , 1991 ; guennoun et al . , 1995 ; coirini et al . , 2003 ) .
moreover , pregnenolone conversion into progesterone has been demonstrated in rat homogenates from septum and amygdala ( weinfeld et al . , 1980 ) .
the co - expression of 3-hsd and gabaa receptor subunits in different brain regions ( laurie et al .
1992 ) gives an anatomo - functional support for the in situ production of progesterone and the gabaa receptor modulation ( guennoun et al .
although regulatory mechanisms underlying the ns biosynthesis inside the brain remain unclear , it is well known the capacity of steroids of negatively modulate the 3-hsd activity in different steroidogenic endocrine glands and in peripheral nervous system , like sciatic nerve ( guennoun et al . , 1995 ; coirini et al . ,
the ru486 caused an impact on the 3-hsd enzyme activity in rat adrenal gland ( albertson et al .
, 1994 ) but not in gonads ( sanchez et al . , 1989 ) . in our work
, we described that ss epoxies caused a dose - dependent decrease on the 3-hsd activity .
in fact , the analogues of pregnanolone produced less inhibition than those with the conformation allopregnanolone - like ( rey et al . , 2013 ) .
ns are endogenous regulators of neuronal excitability ( lambert et al . , 2003 ; akk et al . , 2009 ) .
within the brain , reduced steroids ( like allopregnanolone and pregnanolone ) are highly selective and potent modulators of the gabaa receptor functions ( gasior et al . , 1999 ) .
thus , their anticonvulsant , anesthetics and anxiolytic properties are useful in the treatment of several neurological and psychiatric disorders ( schle et al . , 2011 ) .
neuroprotective effects against adverse early life events ( patchev et al . , 1997 ) and neurogenic effects on neurodegenerative diseases , like ad ( brinton and wang , 2006 ) , have been observed with allopregnanolone administration .
steroids with similar activity like this progesterone metabolite provide big opportunities for therapeutic treatments reducing hormonal side effects ( morrow , 2007 ; reddy , 2010 ) .
the principal disadvantage of endogenous ns administration is their poor bioavailability caused by their rapid in vivo metabolism .
thus , endogenous ns stimulation synthesis or synthetic steroids analogues ( poisbeau et al . , 2014 ) might constitute promising novel strategies for several disorders treatments . the current medicinal chemistry around nas is focused on the development of new ss analogues , having the absolute configuration of natural steroids .
several studies indicate that the 3-hydroxyl configuration is the key for binding and activity , but modifications in the steroid nucleus may emphasize different pharmacophores . among the ss developed are ganaxolone and alpx which have anesthetic and anticonvulsant properties . until now , ganaxolone is the only one ss that has been used on human clinical trials for epilepsy ( nohria et al . , 2010 ) .
on the other hand , the ss ent - neurosteroids produced more potent inhibition of tbps binding from the gabaa receptor than alpx ( qian et al .
. moreover , we found that some ss epoxies reduce the tbps binding and stimulate the flunitrazepam and muscimol binding in a dose - dependent manner ( rey et al . ,
this type of effects was observed with the synthetic allopregnanolone analogue co 2 - 6749 ( gma-839 ; way-141839 ; 3,21-dihydroxy-3-trifluoromethyl-19-nor-5-pregnan-20-one ; vanover et al . , 2000 ) .
in fact , neurosteroidogenic agents , that lack benzodiazepine - like side effects , are promising for the treatment of anxiety and depression ( reddy , 2010 ) .
neuroprotective effects have been described with several ss in hypoxia - induced brain injury models . among others ,
the ss ru486 was able to protect against excitotoxicity and traumatic brain injury ( behl et al .
, 1997 ; mccullers et al . , 2002 ) and the 5-androst-3,5,6-triol showed a neuroprotective action in an ischemic stroke model in vitro ( chen et al . , 2013 ) .
moreover , in perinatal brain tissues submitted to hypoxic conditions , restricted analogues from allopregnanolone or pregnanolone showed similar properties preventing the glial and neuronal damage ( rey et al . , 2013 ) .
on the other hand , neurogenic properties on ad were observed with the 4-estren-3,17-diol and ent - steroid of 17-estradiol administrations ( kousteni et al . , 2002 ; covey , 2009 ) .
another issue to take in consideration for the development of ss is related to the presence of all the enzymes necessary for ns synthesis in the brain ( mensah - nyagan et al .
, 1999 ; agis - balboa et al . , 2006 ; do rego et al . , 2009 ) . although regulatory mechanisms around ns biosynthesis are still unclear , it is well known the capacity of steroids to negatively modulate the 3-hsd activity ( in almost all steroidogenic tissues ) and the importance of a minor effect on these activities by the ss administration . specific enzymes and nuclear hormone receptors for endogenous steroids have structurally defined binding sites .
it is important that the ss should be developed lacking the possibility to bind with high affinity to these proteins .
therefore the ss drugs might not strongly interfere with the natural steroids biosynthesis or their specific receptors
. it would be also advantageous that the half - life of these new ss might be quite different and potentially longer , than those of steroid already used as anticonvulsants , anxiolytics , or another neuroactive - neurogenic agents .
thus , it is likely that the development of new ss for therapeutical use will continue requiring a great deal of effort with the attendant generation of new knowledge .
 neurosteroids , like allopregnanolone and pregnanolone , are endogenous regulators of neuronal excitability . inside the brain , they are highly selective and potent modulators of gabaa receptor activity . 
 their anticonvulsant , anesthetics and anxiolytic properties are useful for the treatments of several neurological and psychiatric disorders via reducing the risks of side effects obtained with the commercial drugs . 
 the principal disadvantages of endogenous neurosteroids administration are their rapid metabolism and their low oral bioavailability . 
 synthetic steroids analogues with major stability or endogenous neurosteroids stimulation synthesis might constitute promising novel strategies for the treatment of several disorders . 
 numerous studies indicate that the 3-hydroxyl configuration is the key for binding and activity , but modifications in the steroid nucleus may emphasize different pharmacophores . 
 so far , several synthetic steroids have been developed with successful neurosteroid - like effects . in this work , 
 we summarize the properties of various synthetic steroids probed in trials throughout the analysis of several neurosteroids - like actions . 
Natural neurosteroids and synthetic steroids
Neurosteroids and GABA
NAS and SS neuroprotective role
Neurosteroids synthesis: steroid effects on 3-HSD activity
Conclusion