Effects of amisulpride on emotional memory using a dual-process model in healthy male volunteers.

Memory dysfunction occurs in a number of neuropsychiatric disorders. Therapeutic psychopharmacological agents may exacerbate such memory impairment. Detailed characterisation of drug-induced memory impairment is therefore important. We recently showed that the D(2)/D(3) antagonist amisulpride quantitatively impairs emotional memory in a randomised placebo-controlled study of 33 healthy volunteers. Current evidence suggests that two qualitatively different processes (recollection and familiarity) contribute to recognition memory and can be investigated using a Dual-Process Signal Detection model. Using such a model, we found that amisulpride levels at encoding were significantly inversely correlated with recollection estimates for emotional but not neutral stimuli or familiarity estimates in healthy male volunteers. This suggests that dopamine antagonism at encoding preferentially impairs the recollection component of emotional memory, relative to the familiarity component. This was supported by receiver operating characteristic analysis. We also found a significantly increased false recognition rate, associated with significantly shorter reaction times for emotional but not neutral stimuli in the amisulpride group. These findings have important implications for our understanding of recognition memory processes, as well as the interpretation of neuropsychological findings in medicated patients.

Clozapine: more than 900 mg/day may be needed.

Patients may fail to respond to clozapine treatment despite use of the maximum licensed UK dosage (900 mg/day) because of ultra-rapid metabolism of the drug. We present the findings of a study of a national clozapine/norclozapine assay service for the period 1997-2005 and three individual case studies of patients treated with clozapine in doses greater than 900 mg/day. Clinicians should be alert to the possibility of treatment failure because of rapid clozapine clearance secondary to genetic factors and heavy cigarette consumption. This may necessitate the use of clozapine in doses up to 1400 mg/day, notably in young male smokers. Doses of greater than 900 mg/day are rarely justified in women. Anyone given relatively high-dose clozapine (600 mg/day or more) should be monitored regularly for adverse events and changes in smoking habit.

Micro-extraction techniques in analytical toxicology: short review.

This paper discusses new developments in plasma micro-extraction techniques in the context of established micro-extraction and protein precipitation methodology. Simple liquid-liquid solvent extraction (LLE) of plasma with direct GC or HPLC analysis of the resulting extract has been used for many years. Butyl acetate and methyl t-butyl ether (MTBE) give efficient extraction of many drugs and metabolites from small volumes of plasma or whole blood at an appropriate pH, and form the upper layer, thus simplifying extract removal. Butyl acetate does not interfere with NPD, ECD or MS in GC, whilst MTBE has a relatively low UV cutoff (220 nm). Thus, HPLC eluents that use a high proportion of an organic component allow MTBE extracts to be analysed directly. 'Salting-out' and extractive derivatization using acetic anhydride or phenylboronic acid can be used with appropriate analytes. As regards protein precipitation, an important consideration is lowering the pH, although this is not feasible with acid-labile analytes. More recent developments include sold-phase micro-extraction (SPME) and liquid-phase micro-extraction (LPME). This latter technique especially may prove invaluable as analytes that cannot easily be extracted with LLE can be isolated simply at low cost with a minimum of apparatus.

Suspected clozapine poisoning in the UK/Eire, 1992-2003.

OBJECTIVE: Toxicological analyses are often performed to investigate suspected poisoning, but the interpretation of results may not be straightforward. We studied suspected poisoning cases 1992-2003 where blood clozapine and N-desmethylclozapine (norclozapine) were measured in order to assess the relationship of these parameters to outcome. METHODS: Samples were referred from clinicians, pathologists/coroners, or via the Clozaril Patient Monitoring Service (CPMS, Novartis). Information was gathered from clinical, post-mortem, or coroners' reports. RESULTS: There were seven fatal [five male, two female; median (range) age 28 (24-41) year] and five non-fatal [four male, one female; median age 35 (26-41) year] clozapine overdoses. The median post-mortem blood clozapine and norclozapine concentrations were 8.2 (3.7-12) and 1.9 (1.4-2.4)mg/L, respectively [median clozapine:norclozapine ratio 4.4 (2.9-5.1)]. The median plasma clozapine and norclozapine concentrations (first or only sample) were 3.9 (1.7-7.0) and 0.40 (0.30-0.70)mg/L, respectively [median clozapine:norclozapine ratio 7.6 (5.3-18)] in the remainder. These overdoses were in patients who were poorly or non-adherent to clozapine, or who had taken tablets prescribed for someone else. In 54 further people who died whilst receiving clozapine [38 male, 16 female; median age 41 (22-70) year], the median post-mortem blood clozapine and norclozapine concentrations were 1.9 (0-7.7, n = 43) and 1.4 (0-6.0, n = 39)mg/L, respectively [median clozapine:norclozapine ratio 1.5 (0.4-7.6, n = 38)]. The median post-mortem increase in blood clozapine and norclozapine as compared to the most recent ante-mortem measurement was 489 (98-5,350)% and 371 (139-831)%, respectively [median sample time before death 14 (0-30, n = 21) days]. CONCLUSION: Clozapine poisoning cannot be diagnosed on the basis of blood clozapine and norclozapine concentrations alone. The analysis of ante-mortem blood specimens collected originally for white cell count monitoring and the blood clozapine:norclozapine ratio may provide additional interpretative information.

Measurement of total mirtazapine and normirtazapine in plasma/serum by liquid chromatography with fluorescence detection.

A simple high performance liquid chromatography (HPLC) method for the measurement of the new antidepressant mirtazapine and its N-demethyl metabolite, normirtazapine, in human plasma or serum during low dose mirtazapine therapy has been developed. A Waters Spherisorb S5 SCX column was used with ammonium perchlorate (50 mmol/l) in methanol/water (95 + 5 (v/v)), apparent pH 6.7, as eluent, and fluorescence detection. Only small volumes of sample (0.2 ml) and extraction solvent are used. An interference study found no significant co-elution with drug or metabolite, although paroxetine co-elutes with the internal standard. The recovery of mirtazapine and normirtazapine (mean +/- S.D.) was 79 +/- 2, and 64 +/- 3%, respectively. The LOD was estimated as 0.5 microg/l, LLOQ was 1 microg/l, with a linear response over the concentration range 4-1000 microg/l (both analytes). The analytes were stable in serum for at least 10 months when stored at -20 degrees C. Intra- and inter-day accuracy were in the range 91-107 and 93-103%, respectively. In clinical samples (n = 14, median mirtazapine dose 45 mg per day, range 15-45 mg per day) the median (range) mirtazapine and normirtazapine concentrations were 26 (8-40) and 21 (8-32) microg/l, respectively.

Adult respiratory distress syndrome and renal failure associated with citalopram overdose.

A 45-year-old man ingested 3000 mg of citalopram hydrobromide (2400 mg citalopram). He presented to the Emergency Department 2 hours post-ingestion with a pulse of 100 beats/min and blood pressure of 120/80 mmHg. His electrocardiogram (ECG) was normal. Chest X-ray showed bilateral shadowing, with no evidence of aspiration of gastric contents. Shortly after, he had three tonic-clonic seizures, requiring intravenous diazepam. Eight hours post-ingestion he became oliguric with deteriorating renal function, despite normal arterial and central venous pressures. He became increasingly hypoxic, with chest X-ray changes compatible with adult respiratory distress syndrome (ARDS). Despite treatment with 100% oxygen and continuous positive airway pressure, his gas exchange continued to deteriorate, requiring intubation and ventilation. His renal function also deteriorated with a peak creatinine of 492 micromol/L on day 4 in the absence of rhabdomyolysis. There was complete spontaneous recovery of renal function after 2 weeks. A peak plasma total citalopram (R+S enantiomers) concentration of 1.92 mg/L was recorded 2 hours post-ingestion. Total norcitalopram concentrations continued to rise up to 24 hours post-ingestion. Citalopram has been associated with seizures, ECG abnormalities, rhabdomyolysis and coma after overdose. The renal and respiratory complications seen in this patient have not been reported previously.

The amelioration of the suffering associated with spinal cord injury with subperception transcranial electrical stimulation.

STUDY DESIGN: Double blind, partial crossover. OBJECTIVES: To evaluate the analgesic activity of a novel cranial electrostimulus in people with spinal cord injury (SCI). SETTING: Hereward College, a residential centre that provides educational facilities for students with disabilities. METHODS: Subjects with SCI experiencing chronic pain were randomly assigned into two groups, one of which received sham and the other transcranial electrostimulation treatment (TCET) on two occasions daily for four successive days. After a 'wash-out' period of 8 weeks all subjects returned and received the identical stimulus that the treated cohort received on the first arm of the study. RESULTS: Pain measurements applied before and after each session indicated that the pain decreased in the treated group to 51% of that reported at the commencement of treatment; reported pain intensity did not decrease significantly in the sham treated subjects. The same (sham) subject group reported experiencing 59% of the pain at the end of the second arm of the study (TCET) as on the first arm (sham). No significant differences were determined between the mood of all subjects estimated before and after each sham or TCET treatment session. The reported analgesic, and combined antidepressant and anxiolytic drug use in subjects receiving TCET on the second arm of the study, was 46% and 53% respectively of the average pre-study drug use. No similar decrease in the use of the drugs was noted in the same subjects after sham treatment on the first arm of the study. Salivary cortisol determinations made prior to and after each sham and treatment session implicated this corticoid in the pain-relieving mode of action of the treatment, but could not be associated with any changes in mood. Subjects receiving TCET had significantly higher urinary 3-methoxy-4-hydroxy-phenylglycol (MHPG) output after the TCET treatment period than sham stimulation, implicating increased central noradrenaline (NA) metabolism in the observed effects. CONCLUSION: The subjects reported less pain during, and immediately after receiving this transcranial treatment, although they were using less medication than when receiving sham treatment.

HPLC of basic drugs on microparticulate strong cation-exchange materials - a review.

Propylsulphonic acid (SCX)-modified silica HPLC columns used with methanol or aqueous methanol eluents of appropriate pH and ionic strength can give good retention and peak shape for basic drugs. In the system studied, eluent pH influenced retention via protonation of basic analytes, the pK(a) of the analyte indicating the pH where retention begins to decrease at constant ionic strength. At constant pH, retention is inversely proportional to ionic strength for protonated bases and quaternary ammonium compounds. The underlying retention mechanism appears to be ion-exchange with the SCX moieties, although ionized surface silanols may also contribute to retention at higher eluent pH values. In capillary electrochromatography (CEC) unprecedented efficiencies, but similar selectivity, to that observed in conventional HPLC have been obtained for a standard range of basic drugs using Waters Spherisorb S3SCX.SCX-modified silica columns can be used in the HPLC of many basic drugs, including some compounds that are poorly retained on unmodified silica using methanol-rich eluents. N-Desalkyl and sulphoxide metabolites are often resolved at an appropriate eluent pH. Even analogues differing by a methylene unit in a side-chain remote from a basic centre are often resolved. Applications of Waters Spherisorb S5SCX columns include HPLC of antimalarials such as chloroquine and quinine, cardioactive drugs, for example amiodarone and flecainide, antipsychotics (clozapine, olanzapine), and antidepressants (amitriptyline, clomipramine, dothiepin, fluoxetine) and their N-desalkyl metabolites. Major practical features of these systems are that (i) acidic and neutral compounds are not retained, (ii) solvent extracts can be injected directly, and (iii) eluent recycling can be performed routinely.

HPLC of sertraline and norsertraline in plasma or serum.

A simple method for the measurement of sertraline and norsertraline in plasma or serum suitable for use in single-dose pharmacokinetic studies has been developed. Internal standard solution, aqueous fenethazine (10 mg/L) (20 microL), and Tris buffer (2 mol/L), pH 10.6) (100 microL) were added to plasma (200 microL). Sertraline, norsertraline and the internal standard were extracted into methyl tert-butyl ether (200 microL) by mixing (30 s) and centrifugation (11,000 r.p.m., 4 min). A portion (100 microL) of the extract was injected onto a Spherisorb S5SCX HPLC column (150 x 4.6 mm i.d.) which was eluted with methanol:water (19 + 1) containing ammonium perchlorate (40 mmol/L), final pH 7.0. Detection was by UV monitoring (215 nm). The concentration of each analyte in each sample was calculated from the calibration graph (peak-height ratio of analyte to that of the internal standard against concentration) obtained after analysis of plasma samples containing known amounts of sertraline and norsertraline. The limit of accurate measurement of the assay was 10 micrograms/L) sertraline and 20 micrograms/L) norsertraline.

Regulation of catabolism of microinjected ribonuclease A. Identification of residues 7-11 as the essential pentapeptide.

We have identified a pentapeptide region of microinjected ribonuclease A that is required for enhanced degradation of this protein during serum withdrawal. We introduced reductively methylated [3H]ribonuclease A, [3H]ribonuclease S-protein (residues 21-124), and [3H]ribonuclease S-peptide (residues 1-20) into the cytosol of human fibroblasts by red cell-mediated microinjection and osmotic lysis of pinosomes. The degradative rates of ribonuclease A and ribonuclease S-peptide are increased 2-fold upon withdrawal of serum, while catabolism of ribonuclease S-protein is not regulated in this manner. Certain fragments of ribonuclease S-peptide are also degraded in a serum-dependent fashion (residues 1-14 and 4-13), while other fragments are not (residues 1-10 and 2-8). [3H]Ribonuclease S-peptide is cleaved into two smaller radioactive peptides during loading into red cell ghosts. We tentatively identified the larger fragment as residues 7-11 based on its molecular weight determined by Sephadex chromatography in the presence of 8 M urea combined with sequential Edman degradation to identify the position of radioactive lysines. The smaller peptide fragment appears to be the amino-terminal dipeptide, Lys-Glu, and/or residues 7-8, Lys-Phe. After microinjection into fibroblasts, the pentapeptide is degraded at an enhanced rate in the absence of serum, while degradation of the dipeptide is not affected. We confirmed that residues 7-11 constitute the larger hydrolysis product of S-peptide by synthesizing this pentapeptide and radiolabeling it by reductive methylation. It migrated at the expected position after Sephadex chromatography in 8 M urea and was further hydrolyzed only slightly during loading into red cells. Finally, degradation of this pentapeptide after injection into fibroblasts was enhanced 2-fold upon serum withdrawal. These results, combined with our other recent studies (McElligott, M. A., Miao, P., and Dice, J. F. (1985) J. Biol. Chem. 260, 11986-11993), suggest that the pentapeptide, Lys-Phe-Glu-Arg-Gln, targets microinjected ribonuclease A to lysosomes for enhanced degradation during serum deprivation.

The assessment of zinc status by the zinc tolerance test in various groups of patients.

The zinc status of young and aged subjects, hypertensives, geriatric patients with leg ulcers and cancer patients has been determined by various means. Serum, saliva, urine and hair zinc were measured by atomic absorption spectrometry and no correlation was observed between the zinc levels in any of these differing matrices. The mean hair and salivary zinc level showed little variation between the differing groups of patients and provided little or no indication of zinc status. The results of the present experiment indicate that the zinc tolerance test, that is, an unequivocal rise in serum zinc following per oral administration of this metal, provides the best indication of zinc status.