Structure Activity Studies of NMDA Receptor Modulation by Neuroactive Steroids

Structure-activity data for steroid modulation of NMDA receptor function indicate that, as is the case for GABAa receptors, small modifications in steroid structure can lead to profound changes in modulatory activity (Table 1). PREGS and all other highly efficacious positive modulators identified to date have a double bond between C-5 and C-6, suggesting that the pregnene structure is important for potentiation (21,58,60). However, reduction of this double bond with the introduction of C-5a stereochemistry yields the pregnane steroid epiallopregnanolone sulfate, which retains moderate potentiating activity (58,59), indicating that the requirement for a double bond is not absolute. In contrast, the sulfated C-5 p isomers, pregnanolone sulfate and epipregnanolone sulfate, are strongly inhibitory, whereas the C-3a, C-5a isomer, allopregnanolone sulfate, shows reduced inhibition of NMDA-induced currents, suggesting that C-5P stereochemistry enhances inhibitory efficacy (21,58-60).

Dehydroepiandrosterone sulfate (DHEAS) is only a weak potentiator, suggesting that the side chain at C-21 is important for potentiation. Similarly, introduction of a ketone or hydroxyl at C-11 or a ketone at C-7 eliminates potentiation (21,58,60). In contrast, there appears to be latitude for modifications to the C-17 side chain, as 20^-dihydro-pregnenolone sulfate and 21-acetoxy-pregnenolone sulfate both retain potentiating activity (21,58,60).

Pregnenolone Sulfate

Epipregnanolone Sulfate

Fig. 2. Space-filling models of pregnenolone sulfate and epipregnanolone sulfate. Space-filling representations of pregnenolone sulfate and epipregnanolone sulfate serve to illustrate the similarity between these two molecules, which have opposite modulatory effects on NMDA receptor function.

The C-3 sulfate is not essential for interaction with the NMDA receptor, as pregnanolone hemisuccinate has inhibitory activity similar to pregnanolone sulfate, and pregnenolone hemisuccinate has potentiating activity similar to PREGS (21,58,60). Nevertheless, a negatively charged group at C-3 appears to be important, as pregnanolone, pregnenolone (PREG), and pregnenolone hemisuccinate methyl ester are all inactive (56,60). A notable exception to this rule is estradiol, which inhibits the NMDA receptor despite having only a hydroxyl group at C-3 (61). It is possible that the reduction in the pKa of the C-3 hydroxyl owing to the aromatic nature of the estradiol A ring allows sufficient ionization for it to function as a negatively charged group. Alternatively, it may be that estradiol derivatives inhibit the NMDA receptor by a different mechanism than pregnane steroids.

Interpretation of structure-activity data is complicated by the recent finding that positive and negative steroid modulators act at separate sites (57-59). Thus, it is possible that a compound could be mistakenly classified as inactive because its positive and negative modulatory effects cancel out. It is possible that competition studies will help to resolve these questions. Alternatively, if the positive and negative steroid modulatory sites on the receptor can be identified, it may be possible to disable one or the other by site-directed mutagenesis to study structure-activity relations of each site separately.

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