Angelos Halaris

Reduced Neuropeptide Y Concentrations in Suicide Brain

Neuropeptide Y (NPY) was measured in postmortem brain tissue from victims of suicide and from individuals dying a sudden natural or accidental death (controls). Concentrations of NPY‐immunoreactivity were measured by radioimmunoassay in frontal cortex (BA 10), temporal cortex (BA 22), caudate nucleus, and cerebellum. Concentrations of NPY‐immunoreactivity were significantly lower in postmortem frontal cortex (−14%) and caudate nucleus (−27%) from suicide victims compared with age‐matched controls. A subgroup of suicides with evidence of a history of depression revealed more robust reductions in concentrations of NPY‐immunoreactivity in frontal cortex and caudate nucleus, as did four individuals who died from natural causes and also were described as having a possible history of depression. Concentrations of NPY‐immunoreactivity in temporal cortex and cerebellum from victims of suicide or from the subgroup of subjects with a possible history of depression were not significantly different from those of age‐matched controls. We suggest there is a deficit in the brain NPY system leading to region‐specific reductions in peptide concentrations in subjects who have a history of depression.

Evaluation of studies on platelet alpha2 adrenoreceptors in depressive illness

Discrepant results have been reported from at least ten laboratories regarding the status of platelet alpha 2 adrenoreceptors in depressed patients. Using a statistical test to combine those studies which utilized radioligand binding techniques, we find the overall data support an elevation in density of platelet alpha 2 adrenoreceptors from drug-free depressed patients (p less than 0.05) and suggest a normalization to lower binding values following antidepressant drug treatment (0.05 less than p less than 0.10). However, these positive results are attributable to highly significant findings by only three laboratories. Much of the discrepancy may be attributable to numerous methodological variables which distinguish the studies. Foremost amongst these variables are the use of different platelet size populations, the use of different medium, and the choice of radioactive ligand and competitor (non-radioactive ligand) in the assay. We present a rationale for the proper choice of each methodological condition used in the clinical assessment of platelet alpha 2 adrenoreceptor status, hoping that improved experimental designs will resolve the current controversy.

Binding of [3H]-p-aminoclonidine to α2adrenoceptor states plus a non-adrenergic site on human platelet plasma membranes

Characterization of the binding of [3H]p-aminoclonidine ([3H]PAC) to purified plasma membranes from human platelets has revealed multiple binding sites. [3H]PAC identified site-1 in the picomolar affinity range (site-1 KD estimates ranged from 13 to 94 pM). Site-1 displayed a rank order of competition by various compounds for [3H]PAC, indicative of an alpha 2-adrenoceptor, and was sensitive to 0.1 mM GTP. [3H]PAC also identified a second site with nanomolar affinity (site-2 KD estimates ranged from 0.7 to 1.7 nM). In the presence of 0.1 mM GTP, site-2 was not diminished significantly. Also in contrast to site-1, site-2 displayed low affinity for yohimbine (YOH), (-)-epinephrine and (-)-norepinephrine (NE). Therefore, site-2 could not be an active alpha 2-adrenoceptor; instead it had properties similar to a previously reported imidazoline-preferring binding site. A third site (site-3) bound [3H]PAC with a KD for site-3 of 26.6 +/- 10.0 nM (SD). Site-3 had a rank order of competition by various compounds for 5 nM [3H]yohimbine ([3H]YOH) binding which was indicative of an alpha 2-adrenoceptor. (-)-NE competed for 5 nM [3H]YOH binding at two sites: site-1 Ki = 32 pM, site-3 Ki = 239 nM. Treatment with 0.1 mM GTP completely removed site-1 and transferred the competitive binding of (-)-NE to low affinity (Ki = 437 nM). Thus, site-3 appears to be a free alpha 2-adrenoceptor. Bmax estimates for untreated membranes, derived from simultaneous multi-experiment curve-fitting analyses, were site-1 = 36 +/- 29 fmol/mg plasma membrane protein, site-2 = 95 +/- 34 fmol/mg and site-3 = 154 +/- 35 fmol/mg. We are the first to report a site for [3H]PAC binding on platelets (site-2) with properties uncharacteristic of an adrenoceptor. This observation appears to be due to our use of purified plasma membrane and low ionic strength buffer. These studies relate to reports of increased binding of [3H]PAC to platelets from depressed patients.

The relationship between life stress and depression in an endogenous sample

The role of stress was examined in a sample of 68 endogenously depressed patients. Antecedent and concurrent life events and long-term difficulties, operationalized by different subcategories and scoring options, were related to depression severity, treatment response versus nonresponse, and maintenance versus relapse. Results showed an association between initial depression and various categories of subjective stress; a reduction in subjective (but not objective) event-related stress coincident with remission; positive relationships between life events and response and maintenance and between difficulties and relapse; and a higher event profile among unipolar depressive patients. Results are discussed in terms of the positive prognostic value of reactivity in endogenous depression, as well as the interactive relationship between continuing depression and long-term difficulties.

Increased imidazoline and α2 adrenergic binding in platelets of women with dysphoric premenstrual syndromes

An association between dysphoric premenstrual syndromes (PMS) and a lifetime history of major depressive disorders has previously been documented. Other studies have demonstrated an increase in the binding of radiolabeled imidazoline compounds to platelets of depressed patients. Clonidine and related imidazoline compounds interact with alpha 2 adrenoceptors to inhibit neuronal noradrenergic activity and in higher concentrations, they stimulate noradrenergic activity through their interaction with imidazoline receptors. Here we report increased 3H para-aminoclonidine binding to high affinity alpha 2 adrenoceptor sites as well as to nonadrenergic imidazoline binding sites in platelets of women with dysphoric PMS. This higher binding was most pronounced during the late-luteal-symptomatic phase of the menstrual cycle and, to a lesser degree, during the non-symptomatic mid-follicular phase. Binding to the imidazoline site distinguished women with dysphoric PMS from women with no such symptoms, was highly positively correlated with the severity of symptoms, and was negatively correlated with plasma levels of progesterone. These findings suggest that platelet imidazoline binding sites might be a biological marker for dysphoric states in PMS or for the vulnerability to develop them. These findings also point to a possible biological link between dysphoric PMS and major depressive disorders.

Chronic desipramine treatment reduces regional neuropeptide Y binding to Y2-type receptors in rat brain

Chronic treatment of rats with desipramine and imipramine (5 mg/kg/twice daily/i.p.) for 14 days caused a significant reduction in the binding of [3H]propionyl NPY to membranes prepared from frontal cortex, nucleus accumbens, hypothalamus and hippocampus. There was no change in binding of [3H]propionyl NPY in the parieto-occipital cortex, striatum or amygdala. Scatchard analysis of binding data from frontal cortical and hippocampal membranes showed that [3H]propionyl NPY bound to a single site with a Kd of approximately 0.3 nM. The loss of [3H]propionyl NPY binding in hippocampal and frontal cortical membranes revealed that chronic tricyclic antidepressant treatment produced a reduction in the number of binding sites with no change in the affinity for the ligand. Chronic desipramine treatment did not alter the ability of NPY (0.01-25 microM) to stimulate inositol phosphate accumulation in rat frontal cortical slices as compared to saline-treated animals. The lack of change of NPY-induced inositol phosphate accumulation following chronic desipramine treatment showed that there was no change to Y1 NPY-type receptors which are linked to the hydrolysis of inositol phospholipids. However, the ability of NPY (0.05-0.5 microM) to inhibit forskolin (1 microM) stimulated adenylate cyclase via Y2 NPY-type receptors in rat frontal cortical slices was significantly reduced following chronic desipramine treatment. This finding suggests that the reduction of [3H]proprionyl NPY binding in selective brain regions may be the result of an antidepressant-induced loss of Y2-type NPY receptors which are negatively linked to adenylate cyclase.

Dimensions of depression: A comparative longitudinal study

A study was conducted to examine the factor structure of a set of commonly employed depression-related measures and to determine whether higher-order composite variables based on factor loadings would be differentially related to changes in depression status. Five cognitive and two behaviorally oriented measures were administered to 66 hospitalized depressives at pretreatment, 4 to 8 weeks later at the end of an initial phase of pharmacological treatment, and at 2- to 4-month intervals during an 8-month follow-up period. A principal-components analysis performed on nine variables produced three interpretable factors that differed systematically with respect to amount of differentiation between responders and nonresponders, as well as magnitude of change associated with successful treatment. Analyses performed on individual measures to track maintainers versus relapsers across three time points likewise yielded results consistent with predictions based on factor membership.

Super high affinity 3H-para-aminoclonidine binding to platelet adrenoceptors in depression

1. An assay was developed using sucrose gradient purified platelet plasma membranes which allowed detection, for the first time in patients, of both super-high affinity (KD = 17 pM) and high affinity (KD = 1.7 nM) binding sites. 2. Limited Scatchard plot analyses were performed on platelet membranes from depressed patients and controls using 10 pM-2.5 nM 3H-p-aminoclonidine (3H-PAC). 3. Patients (n = 9) were age-paired with healthy control subjects for simultaneous blood drawing, platelet preparation and analysis. 4. All patients were endogenous depressives with Hamilton-Depression scores ranging from 19 to 30 at the time of pre-treatment. Seven of the nine patients were analyzed again at six weeks of treatment with antidepressant medication. 5. Using 60 pM 3H-PAC (a concentration determined to bind predominantly to the super-high affinity receptor state) pre-treatment patient values were higher then paired controls (p = 0.06). Post-treatment analysis of seven of the patients and paired controls showed no differences (p = 0.5) suggesting a normalization of receptor binding following treatment. 6. No differences were observed in platelet yield or morphology or in the percent of other blood cell contaminants in the platelet preparations between patients at pre-treatment and controls. However, the platelet yield was significantly lower in patients post-treatment (p = 0.06). 7. These results are in agreement with two previous studies showing elevated 3H-clonidine binding to high affinity sites from depressed patients. The data presented herein suggest that there is a modest 1.25-fold elevated super-high affinity platelet adrenoceptor binding in depressed patients pre-treatment. Receptor binding becomes normal post-treatment.

Diurnal rhythm of 3-methoxy-4-hydroxyphenylglycol (MHPG): relationship between plasma and urinary levels

Plasma and urinary levels of MHPG were determined in six normal volunteers. Samples were obtained at 3-hour intervals for plasma and at 12-hour intervals for urine. Acrophase, amplitude and period were determined for plasma MHPG levels. A sinusoidal pattern was obtained for diurnal plasma MHPG with a peak at 15:00 hrs. +/- 46 min. Urinary MHPG, corrected for creatinine levels, correlated with both 9 AM plasma MHPG and with baseline plasma MHPG. Furthermore, the relationship between plasma and urinary MHPG was linear when the rhythm of urinary levels was assumed to lag 6.2 hours behind the plasma rhythm. It was concluded that free MHPG is evenly distributed in the total body space and that conjugated MHPG is largely restricted to the blood.

A Comparison of the Binding of [3H]Proprionyl‐Neuropeptide Y to Rat and Human Frontal Cortical Membranes

Abstract: The binding characteristics of [3H]proprionyl‐neuropeptide Y ([3H]proprionyl‐NPY) were studied in frontal cortical membranes prepared from rat and human postmortem tissue. The specific binding of NPY decreased as the magnesium concentration increased from 1.05 to 10 mM. The binding was also influenced by the concentration of GTP in the buffer medium, with a resulting 45% decrease in NPY binding in the presence of 10‐‐6M GTP. Using equilibrium binding studies, [3H]proprionyl‐NPY was found to bind in both tissues with high affinity to a single class of receptors with a similar KD (0.035 nM). However, kinetic experiments in both tissues provided evidence for two components of [3H]proprionyl‐NPY binding which may be related to receptor states. Competition binding experiments showed that peptide YY (PYY) was equal to NPY in its ability to displace [3H]proprionyI‐NPY, whereas rat and human pancreatic polypeptide were without effect up to a concentration of 10‐‐6M. This suggests that, whereas PYY and NPY may compete for the same receptor(s), the pancreatic polypeptides probably act on a separate population of receptors.