Plasma melatonin--an index of brain aging in humans?

We investigated the age-related changes in the circadian rhythm of plasma melatonin as a potential index of brain aging in man. The subjects were 5 young men aged 19-25 years, 11 older men aged 51-65 years, 6 elderly men aged 66-89 years, 7 young women aged 19-25 years, 5 premenopausal women aged 45-50 years, 8 postmenopausal women aged 51-65 years, and 5 elderly women aged 66-75 years. They were all physically and psychiatrically normal. Serial blood samples were drawn from 8:00 AM until 8:00 AM on the next day, with the indoor illumination set at 300 Lux from 7:00 AM until 4:00 PM and at 50 Lux thereafter. Plasma melatonin was estimated by radioimmunoassay. The results show that there is a significant negative correlation between age and 24-hr secretion of plasma melatonin (r = -0.952, p less than 0.0001), between age and peak levels of plasma melatonin (r = -0.937, p less than 0.00001), and between age and the lag in time from sunset to the onset of significant elevation of plasma melatonin over daytime values (r = 0.916, p less than 0.0001). It is concluded that study of the circadian rhythm of plasma melatonin may prove to be a useful index of the aging process.

Antagonists of excitatory amino acids and cyclic guanosine monophosphate in cerebellum.

The excitatory amino acid analogues kainate, quisqualate, domoic acid, 4-fluoroglutamate, homocysteic acid and N-methylaspartate as well as the tremor-inducing drugs harmaline and oxotremorine all induced significant elevations in cyclic guanosine monophosphate (cGMP) levels in the cerebellum in vivo. The putative antagonists of excitatory amino acids, 2-amino-5-phosphonovalerate (APV) and piperidine dicarboxylate (PDA) both blocked the actions of the tremorogens. Piperidine dicarboxylate also blocked the in vivo activity of all the amino acid analogues except homocysteic acid and N-methylaspartate. 2-Amino-5-phosphonovalerate (APV) was inactive against kainate, quisqualate and homocysteic acid. It therefore appears that PDA and APV are useful tools for the further study of the function of glutamate and asparatate receptors.

Functionally and anatomically distinct populations of vasopressinergic magnocellular neurons in the female golden hamster.

The present study was done to determine whether the vasopressinergic neurons in the hypothalamus controlling flank marking behavior are distinct from the magnocellular neurons comprising the hypothalamo-neurohypophysial system. Animals were either hypophysectomized or injected with a suicide transport lectin, volkensin, into the neurohypophysis. Both procedures resulted in a pronounced loss of vasopressin-immunoreactive perikarya throughout the hypothalamus concomitant with increases in water intake and urine output and decreases in circulating levels of vasopressin. The loss of the hypothalamo-neurohypophysial system was most pronounced in volkensin-treated animals that presented with frank diabetes insipidus and exceedingly low levels of plasma vasopressin. However, the vasopressinergic fibers and magnocellular neurons in and around the anterior hypothalamus implicated in the control of flank marking survived the volkensin treatment. Volkensin-treated animals exhibited levels of flank marking typical of untreated animals. These data suggest the presence of anatomically and functionally distinct populations of vasopressinergic magnocellular neurons in the hypothalamus of the golden hamster.

Comparative potencies of substance P, substance K and neuromedin K on brain substance P receptors.

The relative potency of substance P, substance K and neuromedin K on [3H]substance P binding has been compared in rat brain membrane preparations. Substance K and neuromedin K, two newly isolated substance P-related mammalian peptides, are very weak competitors for [3H]substance P binding sites. It suggests that these two neuropeptides are not acting on the same receptor type as substance P. Moreover, previous substance P receptor classifications are unlikely to be applicable to brain tachykinin receptor sub-types.

Cytokine indices in Alzheimer's temporal cortex: no changes in mature IL-1 beta or IL-1RA but increases in the associated acute phase proteins IL-6, alpha 2-macroglobulin and C-reactive protein.

Recent immunocytochemical data have demonstrated increases in interleukin-1 beta (IL-1 beta), interleukin-6 (IL-6) and the IL-6-inducible acute phase protein, alpha 2-macroglobulin (alpha 2-M), in Alzheimer's disease (AD) brains. We investigated the levels of these proteins quantitatively using ELISA procedures and determined if increases in IL-1 beta were compensated for by a parallel increase in the endogenous interleukin-1 receptor antagonist (IL-1RA). Comparing control vs. Alzheimer's temporal cortex, we examined mature IL-1 beta, IL-1RA, IL-6, alpha 2-M and C-reactive protein (CRP). The specificities of the ELISA procedures were verified by serial dilutions of the samples; by chromatofocusing, and by Sephadex G-150 gel filtration. There were no differences in the levels of mature IL-1 beta or IL-1RA in AD and control brains. However, IL-6 levels were detectable in 14 of the 16 Alzheimer samples but only 2 of the 14 control samples. There were also significant increases seen in alpha 2-M and CRP levels in the Alzheimer's group compared to controls. These data support previous studies demonstrating a possible up-regulation of neuroimmune function in Alzheimer's cortex; however, we cannot determine, at this time, if this immune reaction is initiated by IL-1 beta.

Phencyclidine binding sites in the nucleus accumbens and phencyclidine-induced hyperactivity are decreased following lesions of the mesolimbic dopamine system.

[3H]Phencyclidine [( 3H]PCP) binding to rat nucleus accumbens, hippocampal and striatal membranes, and PCP-induced locomotor hyperactivity were assessed following selective lesions of the mesolimbic dopaminergic system. 6-Hydroxydopamine (6-OHDA) injections into the A10 region of the ventral tegmental area or into the accumbens itself resulted in a blockade of PCP's stimulatory effects and a highly significant reduction in the number of [3H]PCP binding sites and dopamine content of the nucleus accumbens. However, destruction of the dopaminergic mesolimbic fibers did not significantly alter hippocampal or striatal [3H]PCP binding. The data suggest that PCP elicits its locomotor stimulating effects via an interaction with PCP binding sites located mostly on mesolimbic dopaminergic terminals within the nucleus accumbens.

Autoradiographic distribution of [3H]dexoxadrol (a phencyclidine-related ligand) binding sites in rat and human brain.

[3H]dexoxadrol, a dissociative anesthetic, binds with high affinity to specific sites in rat brain (membrane binding and light microscopic autoradiography). Various phencyclidine (PCP) analogues compete for [3H]dexoxadrol sites in a slightly different manner than against [3H]PCP binding sites. As for [3H]PCP binding, [3H]dexoxadrol binding sites are highly concentrated in brain regions such as the cortex and the hippocampus. However, other areas such as the hypothalamus are enriched only in [3H]dexoxadrol binding sites. This suggests that [3H]dexoxadrol binds to PCP-related sites in certain brain regions but not in others. In the human forebrain, [3H]dexoxadrol binding sites are distributed as in the rat brain and mainly found in the caudate, putamen and cortex.

Circadian rhythm of plasma melatonin in endogenous depression.

The circadian rhythm of plasma melatonin was investigated in normal men 18-30 years (N = 5), normal men 50-70 years (N = 5) and in six patients with endogenous depression. The environmental photoperiod was 11 hours. The subjects and patients were indoors with lights on from 07:00 until 23:00 hours. Blood samples were obtained every 4 hours over a 24 hour period, with additional sampling at 22:00 and 02:00 hours. Plasma melatonin was estimated by radioimmunoassay compared to both groups of controls. In the depressed patients, the levels of melatonin were low throughout the 24 hour period. The depressives had a delayed onset of the dark phase of the rhythm. The patients also showed peak melatonin levels occurring earlier than in the controls. Circadian rhythm of melatonin and therefore of its pacemaker may be altered in endogenous depression.

Effect of apomorphine on melatonin secretion in normal subjects.

There is some evidence in animals that dopamine (DA) affects melatonin secretion. The effect of apomorphine (Apo), a selective DA receptor agonist, and placebo on day-time melatonin secretion was studied in six normal men. Apo HCl in a dose (0.5 mg sc) which increased growth hormone secretion in all subjects had no effect on day-time melatonin concentrations in plasma. In keeping with other clinical studies these data suggest that melatonin secretion is not regulated by a DA mechanism in man.

Autoradiographic distribution of multiple classes of opioid receptor binding sites in human forebrain.

Receptor binding parameters and autoradiographic distribution of various opioid receptor sites have been investigated in normal human brain, post-mortem. [3H]DAGO, a highly selective mu ligand, binds to a single class of high affinity (Kd = 1.1 nM), low capacity (Bmax = 160 fmol/mg protein) sites in membrane preparations of frontal cortex. These sites show a ligand selectivity profile that resembles that of the mu opioid receptor. On the other hand, [3H]bremazocine, in presence of saturating concentrations of mu and delta blockers, appears to selectively bind to a single population of kappa opioid sites (Kd = 0.13 nM; Bmax = 93.0 fmol/mg protein) in human frontal cortex. Whole hemisphere in vitro receptor autoradiography reveals that [3H]DAGO-mu, [3H]DSLET-delta and [3H]bremazocine (plus blockers)-kappa binding sites are discretely and differentially distributed in human forebrain. In the cortex, mu sites are concentrated in laminae I and IV, delta sites in laminae I and II while kappa sites are found in deeper layers (laminae V and VI). In subcortical nuclei, high densities of mu and delta sites are seen in the caudate and putamen while high amounts of kappa sites are present in the claustrum and amygdala. The nucleus basalis of Meynert is enriched in all three classes of sites while the globus pallidus only contains moderate densities of kappa sites. Thus, the possible alterations of these various classes of opioid receptors in neurological and psychiatric diseases certainly deserve further investigation.

[3H]-SKF 10047 binding to rat brain membranes: evidence for kappa isoreceptors.

Studies of the binding of [3H]-SKF10047 to rat brain membrane preparations revealed a binding site which possessed high affinity for kappa and agonist/antagonist analgesics. However, this site was distinguished from kappa sites labelled with [3H]-ethylketazocine by its high affinity for Met5-enkephalin Arg6.Phe7 and beta-endorphin as well as its low affinity for dynorphin 1-13. We therefore tentatively suggest that this may represent a possible kappa isoreceptor population.

Effect of imipramine on the circadian rhythm of plasma melatonin in unipolar depression.

The authors took a series of 20 serum samples over a 24-hr period for measurement of melatonin in four men and six women with Major Depressive Disorder (DSM-III), at baseline (depressed) and after 4 weeks on imipramine (150-200 mg/day) after achieving clinical remission. After successful treatment with imipramine, 24-hr secretion and peak levels of melatonin were significantly higher than at baseline, with no difference in time of peak level. Testing after 2 weeks of treatment (four subjects), with only a partial or no clinical response, revealed no differences compared with baseline. The therapeutic efficacy of imipramine may be associated with an enhancement of noradrenergic activity.

Interleukin-1beta and tumor necrosis factor alpha inhibit chondrogenesis by human mesenchymal stem cells through NF-kappaB-dependent pathways.

OBJECTIVE: The differentiation of mesenchymal stem cells (MSCs) into chondrocytes provides an attractive basis for the repair and regeneration of articular cartilage. Under clinical conditions, chondrogenesis will often need to occur in the presence of mediators of inflammation produced in response to injury or disease. The purpose of this study was to examine the effects of 2 important inflammatory cytokines, interleukin-1beta (IL-1beta) and tumor necrosis factor alpha (TNFalpha), on the chondrogenic behavior of human MSCs. METHODS: Aggregate cultures of MSCs recovered from the femoral intermedullary canal were used. Chondrogenesis was assessed by the expression of relevant transcripts by quantitative reverse transcription-polymerase chain reaction analysis and examination of aggregates by histologic and immunohistochemical analyses. The possible involvement of NF-kappaB in mediating the effects of IL-1beta was examined by delivering a luciferase reporter construct and a dominant-negative inhibitor of NF-kappaB (suppressor-repressor form of IkappaB [srIkappaB]) with adenovirus vectors. RESULTS: Both IL-1beta and TNFalpha inhibited chondrogenesis in a dose-dependent manner. This was associated with a marked activation of NF-kappaB. Delivery of srIkappaB abrogated the activation of NF-kappaB and rescued the chondrogenic response. Although expression of type X collagen followed this pattern, other markers of hypertrophic differentiation responded differently. Matrix metalloproteinase 13 was induced by IL-1beta in a NF-kappaB-dependent manner. Alkaline phosphatase activity, in contrast, was inhibited by IL-1beta regardless of srIkappaB delivery. CONCLUSION: Cell-based repair of lesions in articular cartilage will be compromised in inflamed joints. Strategies for enabling repair under these conditions include the use of specific antagonists of individual pyrogens, such as IL-1beta and TNFalpha, or the targeting of important intracellular mediators, such as NF-kappaB.

Localization of kappa opioid receptor binding sites in human forebrain using [3H]U69,593: comparison with [3H]bremazocine.

1. The autoradiographic distribution of kappa opioid receptor binding sites in human brain was examined using two radiolabeled probes, namely [3H]U69,593 and [3H]bremazocine. 2. [3H]U69,593 binding was performed in the absence of blockers for other sites, while [3H]bremazocine binding was investigated in the presence of saturating concentrations of mu and delta blockers to ensure selective labeling of kappa opioid receptors. 3. Our results show that the autoradiographic distribution of [3H]U69,593 and [3H]bremazocine (plus blockers) binding sites is identical, with high densities of sites found in deep cortical layers and claustrum. 4. This indicates that [3H]U69,593 is a highly selective ligand of the kappa opioid receptor type.

Mu opioid receptor binding sites in human brain.

Our experiments focused on the examination of the distribution of mu opioid receptor binding sites in normal human brain using the highly selective ligand [3H]DAGO, in both membrane binding assay and in vitro receptor autoradiography. Mu opioid binding sites are very discretely distributed in human brain with high densities of sites found in the posterior amygdala, caudate, putamen, hypothalamus and certain cortical areas. Moreover the autoradiographic distribution of [3H]DAGO binding sites clearly reveals the discrete lamination (layers I and III-IV) of mu sites in cortical areas.

Nerve growth factor receptors are preaggregated and immobile on responsive cells.

It has been hypothesized that signal transduction occurs by ligand-induced receptor clustering and immobilization. For many peptide receptors, cross-linking by anti-receptor antibodies is sufficient for receptor activation. This is not, however, the case for nerve growth factor receptor (NGFR). Using fluorescence microscopy and fluorescence recovery after photobleaching (FRAP), we have analyzed the distribution and diffusibility of NGFR on a series of cell lines. We have found the following: (1) Cells expressing high-affinity responsive NGFR's display clustered NGFR's even in the absence of ligand. In contrast, NGFR's in nonresponsive cell lines are diffusely distributed. (2) Receptors on responsive cell lines are largely nondiffusing while most receptors on nonresponsive cell lines are relatively free to diffuse. (3) NGF does not greatly alter the distribution or diffusion properties of the NGFR on either nonresponsive or responsive cell lines. Thus, NGFR is preclustered and immobile on responsive cells, which suggests that immobilization of NGFR prior to ligand binding is required for signal transduction.

Unique [3H]tryptamine binding sites in rat brain: distribution and pharmacology.

[3H]Tryptamine binding in rat brain is widely distributed with highest densities in the cortex, striatum and hippocampus. This binding is stereospecific and is potently displaced by tryptamine analogues and beta-carbolines. Phenethylamines also possess a weaker activity in displacing [3H]tryptamine. The strict structural requirements for binding to this site substantiate its unique character and suggest that it may represent a tryptamine receptor in the CNS.

WIN 44,441: A Stereospecific and Long-Acting Narcotic Antagonist.

The opiate antagonist WIN 44,441-3 is a potent, stereospecific antagonist of mu, delta, and kappa opiate receptors. This antagonist activity is of long duration (> 4 h) with no agonist activity being observed. It therefore appears that WIN 44,441-3 will be a useful long-acting opiate antagonist for in vivo studies.