Subjects

The study population consisted of 24 healthy male volunteers aged between 18 and 40, recruited by advertisement from the local population. All were right-handed as assessed using Briggs' modification of the handedness inventory of Annett (1967) (Briggs and Nebes 1975). The inclusion criteria required that subjects had an IQ of 90 or more as assessed by the National Adult Reading Test (NART) and be fluent in English to be familiar with all the words used in the experiment. Subjects were excluded if they had any significant past or current medical history, or any personal or first-degree family history of psychiatric illness. Baseline mood was assessed using the Beck Depression Inventory (BDI) (Beck et al. 1961), and subjects were not included if they scored 8 or more. They were required not to be taking any medication with the exception of paracetamol (acetaminophen). Subjects provided written informed consent prior to participation, and they were reimbursed for their time and expenses. Ethical approval was obtained from the Newcastle and North Tyneside Local Research Ethics Committee.

Experimental design

A double-blind placebo-controlled crossover design was used. Electroencephalographic (EEG) recordings were made from each subject during two separate visits following a 7-day course of 150 mg DHEA, or placebo, twice daily (i.e. a total daily dose of 300 mg). The treatments were administered in a random, balanced order with at least a 4-week interval between treatment periods to exclude any carry-over effects of DHEA and minimise the learning effect of the memory test. Subjects were asked to record the time they took medication and the duration and quality of sleep in a logbook.

Participants attended the Department of Psychiatry, the Royal Victoria Infirmary (RVI), Newcastle upon Tyne at 0850 hours. They were given breakfast and decaffeinated tea or coffee. The last dose of treatment was administered at 0900 hours, followed by the placement of an electrode cap on the scalp for EEG recordings. Visual analogue scale (VAS) measures were administered, and the subjects were requested to report any adverse and/or beneficial effect of treatment they may have noticed during the last week. The purpose of the experiment and the instructions were explained to the subjects thoroughly.

Dehydroepiandrosterone and cortisol assay

Four saliva samples were collected 1 day prior to each visit at 1200, 1600, 2100 (just before the evening dose of medication) and 2200 hours. A further five saliva samples were collected on the day of testing at 0900 (baseline before last dose of medication), 0930, 1000, 1100 and 1230 hours. Samples were collected by passive drooling (spitting into a plastic tube), without using aids to salivation or swabs. Cortisol and DHEA concentrations in the saliva samples were assayed using a coated tube radio-immunoassay (RIA) kit obtained from M P Biomedicals (Tyne & Wear, UK). Intra-assay variations for cortisol and DHEA were 6.2 and 8.3%, and inter-assay variations 3.0 and 4.2%, respectively.

Visual analogue scale

VASs were used to assess subjective feelings of mood, well-being, memory, sexual drive, appetite and alertness. The VAS measures consisted of a 10-cm bar with ‘best’ and ‘worst’ indicated at its extremities for each variable.

Experimental items for event-related potential procedure

These were identical to material employed in previous studies (Wilding and Rugg 1996; McAllister-Williams and Rugg 2002). In brief, stimuli consisted of low frequency (1–7 per million) words selected from Kucera and Francis corpus (1967). In the study phase, subjects were presented with two lists of words presented binaurally. In each word list, half of the words were spoken in a male voice and half in a female voice, randomly determined. Associated test lists were created with 50% old words presented in the study lists and 50% new words. Test lists of words were presented visually on a computer monitor, with each word presented for 500 ms and subtending a vertical angle of 0.5° and a maximum horizontal angle of 2.8°. Subjects were exposed to two different study/test lists on each of the two recording sessions.

Episodic memory task

Subjects were informed that the aim of the experiment was to investigate memory for spoken words. On each of the two visits, subjects underwent an orientation and preliminary practice session utilising study and test words not included in the actual experiment. Following the practice, subjects undertook two study/test cycles, as described above.

As in previous investigations (Wilding and Rugg 1996; McAllister-Williams and Rugg 2002), the voice in which each study item was presented dictated which of the two encoding tasks should be performed. Subjects were instructed to listen to each word and to respond verbally by repeating the word aloud and then judge whether it was active/passive or pleasant/unpleasant. This procedure was performed to enhance the encoding process. The mapping of task to gender was counterbalanced across subjects.

The study phase was followed by a period of 15 min rest, during which the subjects' attention was distracted and then the test phase was conducted. First, an asterisk appeared on the screen for 1 s as a fixation point and to advise the subjects that they were about to see the stimulus word. Then a word was presented, and the subjects were asked to respond as quickly and accurately as possible as to whether this was an old word they had heard during the study phase or a new one, using the thumb of either their left or right hand. A question mark appeared on the screen following the subjects' response for 2.5 s, and they were instructed that when they see if the word was old they should indicate the gender of the voice that spoke the word and respond by pressing one of the two buttons. No response was required if the word was new. For each subject, the same evaluation of the voice (pleasant/unpleasant or active/passive) and the same button assignment (old/new, male/female) remained consistent in both visits to avoid any possible confusion. These voice and button assignments were counterbalanced across subjects to ensure that there was no correlation between the hands used for old/new and male/female judgement. The total time including orientation/practice study-test block and two experimental study-test blocks was approximately 75 min.

Event-related potential recording

EEG was recorded using an elasticated cap (Easy Caps, Germany) with 29 silver/silver chloride electrodes placed on the scalp in accordance with the International 10–20 system (American Electroencephalographic Society 1994). Two additional electrodes were placed on mastoid processes, with the left mastoid electrode as a reference to all channels, and ERPs were algebraically reconstructed off-line to represent recordings with respect to an average mastoid reference. Vertical electrooculogram (EOG) was recorded between electrodes placed below each eye and an electrode placed on the nasion. Horizontal EOG was recorded between electrodes placed on the outer canthus of the left and right eyes. EEG and EOG were filtered with a bandpass of 0.01–100 Hz and sampled at a rate of 6 ms per point for an epoch of 1536 ms beginning 102 ms before the onset of words presented in the test phase.

Average ERPs were generated for each subject for recognised old words attracting correct source judgements and for correctly identified new items. To maximise the number of trials available for averaging, a blink-correction procedure was employed utilising vertical EOG recordings. Any trial containing residual artefact was rejected if any channel, except vertical EOG, had a voltage deflection greater than ±75 μv. To maintain an acceptable signal/noise ratio, a lower limit of 20 artefact-free trials per subject per visit per response category was set.

Source localisation of the electric activity

LORETA, a source localisation technique, was used to estimate the three-dimensional intracerebral current density distribution from the scalp electric potential differences (Pascual-Marqui et al. 1994, 1999). In this method, the cortex is modelled as 2394 voxels using the digitised Talairach atlas (Talairach and Tournoux 1988), with a spatial resolution of 0.343 cm3 (Pascual-Marqui et al. 1999). LORETA depends on a smoothness assumption according to which neighbouring neuronal populations show highly correlated activity, thus solving the non-unique ‘inverse’ problem that results from the calculation of the electric sources from potentials recorded on the scalp surface (Pascual-Marqui et al. 1999). The resulting solution has relatively low spatial resolution, preserving the location of maximal activation but with some dispersion. In recent years, accumulating literature has shown LORETA localisation to be consistent with functional magnetic resonance imaging (fMRI) results (Seeck et al. 1998). However, the validity of LORETA solutions, particularly localisation of small and deep electrical generators such as the hippocampus, has been questioned (Grave de Peralta Menendez et al. 2000; Phillips et al. 2002; Fontanarosa et al. 2004). As a consequence, the results of LORETA in this study were treated with caution and simply an adjunct to topographical analysis of the ERP data.

Statistical analysis

All values are quoted as means±standard deviations. Statistical comparisons were made using analysis of variance (ANOVA) incorporating the Geisser–Greenhouse correction for inhomogeneity of covariance. F ratios are reported with corrected degrees of freedom. Statistical significance was adjudged at the p<0.05 level.

LORETA software (LORETA-KEY version June 2003; The Key Institute for Brain-Mind Research, Switzerland) was used to perform statistical non-parametric mapping (Pascual-Marqui et al. 1994, 1999). To identify time periods of statistical difference between ERP scalp maps associated with different conditions, topographic analysis of variance (TANOVA) was conducted to calculate the probability of dissimilarity for each response at 6 ms intervals from −102 to 1434 ms relative to stimulus presentation. This procedure is a non-parametric randomisation test computing statistical significance for each pair of maps, correcting for multiple comparison (Thomas and Holmes 2002). Following identification of the statistically significant differences in scalp activity by TANOVA, LORETA was utilised to identify underlying neural generators during the same time period. A LORETA image was generated within the significant time period for each cortical voxel. Statistical non-parametric paired t tests were performed for the comparison of current density distribution between conditions on a voxel-by-voxel basis, corrected for multiple testing.