Delivery of peptides and proteins through the blood-brain barrier.

Peptide and protein therapeutics are generally excluded from transport from blood to brain, owing to the negligible permeability of these drugs to the brain capillary endothelial wall, which makes up the blood-brain barrier (BBB) in vivo. However, peptides or protein therapeutics may be delivered to the brain with the use of the chimeric peptide strategy for peptide drug delivery. Chimeric peptides are formed when a non-transportable peptide therapeutic is coupled to a BBB drug transport vector. Transport vectors are proteins such as cationized albumin, or the OX26 monoclonal antibody to the transferrin receptor; these proteins undergo absorptive-mediated and receptor-mediated transcytosis through the BBB, respectively. In addition to vector development, another important element of the chimeric peptide strategy is the design of strategies for coupling drugs to the vector that give high efficiency coupling and result in the liberation of biologically active peptides following cleavage of the bond linking the therapeutic and the transport vector. The avidin/biotin system has been recently shown to be advantageous in fulfilling these criteria for successful linker strategies. The use of the OX26 monoclonal antibody, the use of the avidin/biotin system as a linker strategy, and the design of a vasoactive intestinal peptide (VIP) analogue that is suitable for monobiotinylation and retention of biologic activity following cleavage, allowed for the recent demonstration of in vivo pharmacologic effects in brain following the systemic administration of relatively low doses (12 microg/kg) of neuropeptide.

The melanocortin melanocyte-stimulating hormone/adrenocorticotropin(4-10) decreases body fat in humans.

The control of body fat is a prominent factor in human health. Animal studies have indicated a homeostatic central nervous system regulation of body fat with particular involvement of the melanocortin receptor pathway. This study provides evidence for a similar role for melanocortins in the long-term control of fat stores in humans. Thirty-six normal weight humans were assigned to one of three experimental groups. After a 4-week baseline, one group was treated with MSH/ACTH(4-10) (MSH/ACTH(4-10)) representing the core sequence of all melanocortins. Another group received desacetyl-alphaMSH, a selective agonist of the brain melanocortin-4 receptor, which shares the 4-10 sequence with MSH/ACTH(4-10). The third group received placebo. Treatments were given intranasally twice daily for 6 weeks, at equimolar doses (MSH/ACTH(4-10), 0.5 mg; desacetyl-alphaMSH, 0.84 mg). Body weight, body composition, and plasma hormone concentrations were measured before and after treatment. MSH/ACTH(4-10) reduced body fat, on the average, by 1.68 kg (P < 0.05) and body weight by 0.79 kg (P < 0.001). Concurrently, plasma leptin levels were decreased by 24% (P < 0.02), and insulin levels were decreased by 20% (P< 0.05) after MSH/ACTH(4-10). Changes after desacetyl-alphaMSH remained nonsignificant. The finding of reduced body adiposity after MSH/ACTH(4-10) confirms and extends to the human the findings of animal models indicating an essential role of the hypothalamic melanocortin system in body weight control.

The O-demethylation of the antidementia drug galanthamine is catalysed by cytochrome P450 2D6.

Galanthamine proved effective in symptomatic treatment of senile dementia of Alzheimer's type. The aim of this study was to elucidate the metabolism of galanthamine. Two novel metabolites of galanthamine have been isolated from the urine of eight young men after single doses of 10-15 mg. Some 19.8% of the doses were excreted as O-demethylgalanthamine glucuronide, 5% as N-demethylgalanthamine, 25.1% as galanthamine, and 0.8% as epigalanthamine. After coadministration of quinidine hydrogen sulfate, which inhibits cytochrome P450 2D6 (CYP2D6) selectively, O-demethylgalanthamine glucuronide was highly diminished in urine. In vitro, human liver microsomes metabolized galanthamine to O-demethylgalanthamine with Vmax 5.2 nmol/mg protein/h and Km 187 microM. Ki of quinidine to inhibit O-demethylation was 28 nM. To inhibit cholinesterases, O-demethylgalanthamine was 10-fold more selective for acetylcholinesterase (AChE) versus butyrylcholinesterase (BuChE) than galanthamine. After glucuronidation, O-demethylgalanthamine failed to inhibit AChE and BuChE. N-Demethylgalanthamine inhibited cholinesterases less potently than galanthamine.

Pharmacokinetics of galanthamine in humans and corresponding cholinesterase inhibition.

Measurements were done to determine the plasma concentrations of galanthamine and two of its metabolites, as well as the corresponding inhibition of acetylcholinesterase activity in erythrocytes after applying 5 and 10 mg galanthamine hydrobromide as a constant-rate intravenous infusion for 30 minutes and single oral doses of 10 mg in eight healthy male volunteers. The data obtained revealed first-order pharmacokinetics, complete oral bioavailability, and a mean terminal half-life of 5.68 hours (95% confidence interval, 5.17 to 6.25 hours). Renal clearance accounted for only 25% of the total plasma clearance (CL = 0.34 L.kg-1.hr-1). Only negligible quantities of the putative metabolites, epigalanthamine and galanthaminone, were detected in blood and urine. The inhibition of acetylcholinesterase activity was closely correlated with the pharmacokinetics of galanthamine, a median maximal value of 53% being achieved by applying 10 mg galanthamine intravenously. Analysis of in vitro and ex vivo concentration responses revealed no differences, indicating that no metabolites of galanthamine exert additional inhibition of acetylcholinesterase activity.

Does short-term treatment with modafinil affect blood pressure in patients with obstructive sleep apnea?

OBJECTIVE: To investigate the effects of modafinil, a central nonamphetamine awakening substance, on blood pressure and heart rate in hypersomnolent patients with obstructive sleep apnea. DESIGN: This double-blind, randomized, placebo-controlled crossover trial was performed over 2 days and 3 nights in a single-center study of hospitalized patients from a referred care center. Twenty-six otherwise healthy men (age range, 30 to 60 years) with mild to moderate obstructive sleep apnea were recruited by the outpatient department of the Marburg University Sleep Laboratory. Patients were given 200 mg oral modafinil in the morning and 100 mg at midday. Placebo was given in the same manner in a crossover design. Mean arterial (radial) blood pressure was monitored continuously during nocturnal sleep and during a series of standardized daytime physical and psychologic performance tests. RESULTS: The difference in the main end point between the treatment with modafinil and placebo was 1.17+/-0.83 (mean +/- SE) mm Hg (95% confidence interval: -0.56 to 2.91 mm Hg). The maximal differences in blood pressure values occurred under loaded conditions (systolic blood pressure, ergometry: 5.62+/-1.13 mm Hg; mental stress test: 6.19+/-1.33 mm Hg). CONCLUSION: Short-term administration of modafinil did not elicit a significant response with regard to the main end point. However, cardiovascular effects during mental and physical load were observed. Longterm studies that include subjects with hypertension are necessary to investigate the clinical relevance of the cardiovascular effects of modafinil.

Galanthamine: pharmacokinetics, tissue distribution and cholinesterase inhibition in brain of mice.

Galanthamine was determined in plasma and tissue extracts of mice, after the application of 4, 6 and 8 mg/kg (i.v.), by reverse phase HPLC, with fluorescence detection. A biexponential decline of concentrations in plasma, with a terminal half-life of 43.3 min, was observed after the dose of 4 mg/kg. The volume of distribution (Vss) of 2.17 l/kg was similar to that found in other species, including man. Metabolism to the inactive diastereomer, epigalanthamine, was very limited. There was a rapid accumulation of galanthamine in tissues, which was most pronounced in the kidney (10-fold compared to plasma) and liver (5-fold). In brain, accumulation was similar to other parenchymatous organs (diaphragm, lung) and amounted to 2.10-fold. Red blood cells showed a concentration 1.34-fold greater than plasma. The accumulation of galanthamine in tissue, with the exception of liver and kidney, can be explained by passive distribution according to differences in pH, between intra- and extracellular compartments. Extraction of galanthamine from blood to brain tissue was complete, indicated by a clearance in the range of cerebral blood flow (1.05 ml min-1 g-1). The concentration-time course of galanthamine in brain tissue was parallel to that in plasma during the terminal elimination phase. Measurement of inhibition of acetylcholinesterase (AChE) in the same samples from brain revealed a maximum apparent inhibition of 43% in the homogenate of brain (1:4 w/v in phosphate buffer, 4 mg/kg, 5 min after injection).(ABSTRACT TRUNCATED AT 250 WORDS)

No increase in blood-brain barrier permeability after intraperitoneal injection of endotoxin in the rat.

Reactions mediated by the brain are part of the response to intraperitoneal administration of endotoxin, a model of gram-negative bacterial infection. To test the hypothesis that a compromised blood-brain barrier (BBB) may contribute to these reactions, the integrity of the BBB was measured following lipopolysaccharide administration. Rats received intraperitoneal injections of 50 microg/kg or 2 mg/kg of endotoxin. Brain uptake of a macromolecular vascular marker, 3H-labelled rat serum albumin, and of a poorly permeable low molecular weight substance, [14C]sucrose, was then measured with the intravenous bolus injection method. Compared to controls, neither dose of endotoxin affected the BBB permeability for these tracers. This was true when brain uptake was measured 5 min or 2 h after lipopolysaccharide injection. It is concluded that intraperitoneal endotoxin even at a high dose does not acutely disrupt the BBB.

Acute endocrine failure after brain death?

After brain death, 32 potential organ donors were studied to determine serum and plasma concentrations of hypothalamic-pituitary hormones, thyroid hormones, and cortisol over a period of up to 80 hr. Diagnosis of brain death was established either on the basis of clinical criteria (n = 16) or by angiography (n = 16). While 78% of the organ donors developed diabetes insipidus, none of the circulating hormones of the anterior pituitary gland showed a progressive decline in concentration according to their plasma half-lives. With the exception of arginine vasopressin (AVP), no hormone concentration was found to be subnormal due to the onset of brain death. The subnormal free triiodothyronine (FT3) values in 62% of cases (median FT3 of 2.2 pmol/L within the first 24 hr) and the cortisol concentration of 6.9 micrograms/dl correlate with the frequency of similar findings in patients with severe head injuries. While the adrenocorticotropic hormone (ACTH) concentrations of 10-53 pg/ml remained constant during the study period, thyroid-stimulating hormone (TSH) and human growth hormone (hGH) concentrations showed a 12- and 35-fold increase from baseline values after 30-40 hr. These results suggest that, despite the now generally accepted criteria of brain death, there is still some residual function, and thus also perfusion of the hypothalamic-pituitary neuroendocrine system. This residual function appears to be sufficient to maintain hormonal plasma levels at least in the low reference range in most donors. Hormonal depletion in organ donors subsequent to brain death, as suggested repeatedly in the literature, could not be confirmed. The analysis of serum or plasma concentration patterns of a number of hormonal parameters following brain death does not support the rationale for a routine replacement therapy of total triiodothyronine (TT3) or cortisol to maintain endocrine homeostasis prior to organ harvest. However, dexamethasone therapy may be followed by suppression of the adrenal cortex of the organ donor. In these cases, cortisol substitution may be indicated.

In vivo demonstration of subcellular localization of anti-transferrin receptor monoclonal antibody-colloidal gold conjugate in brain capillary endothelium.

To study transcytosis at the blood-brain barrier (BBB) in vivo, we used the internal carotid artery perfusion technique in rats. Brain uptake of the OX26 anti-transferrin receptor antibody (IgG2a) coupled to 5-nm colloidal gold (OX26-Au) was morphologically examined after infusion or perfusion into the carotid artery for 10 min. The brain tissue was subsequently perfusion-fixed with 2% glutaraldehyde. By light microscopy, silver enhancement of vibratome sections revealed staining of the vascular tree. Electron microscopy showed binding of gold particles at the luminal plasma membrane of brain capillary endothelia, endocytosis in vesicles (50-100 nm), and particles beyond the abluminal plasma membrane. Studies were performed with an IgG2a isotype control, UPC10. Internal carotid artery infusion of [125I]-UPC10 showed no evidence of brain uptake or binding to endothelium. However, microvessel structures were identified after silver enhancement of vibratome sections of brain following internal carotid artery infusion of 5-nm colloidal gold conjugates of UPC10 (UPC10-Au). The morphologically observed binding of UPC10-Au to brain microvessels may be induced by gold conjugation. These studies describe the use of gold conjugates of antibodies to delineate the subcellular pathway involved in transcytosis through the BBB.

Cationization of a monoclonal antibody to the human immunodeficiency virus REV protein enhances cellular uptake but does not impair antigen binding of the antibody.

Replication of the human immunodeficiency virus (HIV) within cells may be blocked by neutralization of viral-specific proteins that are absolutely required for growth of the virus. One such viral-specific protein is REV, and a monoclonal antibody (mAb) against the REV protein is a potential therapeutic for acquired immune deficiency syndrome (AIDS). However, in order to effect 'intracellular immunization', mAbs must be enabled to target the intracellular compartment. One strategy for transcellular drug delivery of mAb-based therapeutics is cationization, and the present studies describe the cationization of a murine mAb specific to the REV protein of HIV-1. The isoelectric point (pI) of the mAb was raised from 6.6 to more than 9.5. There was virtually no difference in binding to wild-type REV protein between the native or cationized anti-REV mAb, based on studies with a solid-phase immunoradiometric assay. The uptake of the [125I] native anti-REV mAb by human peripheral blood lymphocytes (PBLs) was negligible; however, there was a marked increase in both total cell binding and endocytosis by the human PBLs of the [125I] cationized anti-REV mAb. In conclusion, these studies show that an anti-REV mAb may be cationized to markedly increase endocytosis of the antibody and that this cationization reaction does not significantly alter the affinity of the antibody for its target protein. Cationized anti-REV mAbs may allow for intracellular immunization of the virus and are potential therapeutics for the treatment of HIV.

Hormonal responses to exhausting physical exercise: the role of predictability and controllability of the situation.

Psychological conditions which produce sustained activation have been clearly identified. Among these are the predictability and the controllability of the situation. We studied the impact of these psychological variables on hormone secretion (cortisol, ACTH, vasopressin, prolactin, and hGH plasma levels) under a standardized physical load. Sixteen subjects participated in four sessions each, one week apart, with the task of riding a bicycle until exhaustion. During three sessions, all experimental conditions were held identical to ensure the situation was a predictable as possible. During the fourth session, instructions induced a certain level of uncontrollability. Whereas physiological and performance measures did not vary with experience in the task, cortisol, ACTH, and vasopressin responses declined with increasing experience. This emphasizes the importance of the psychological definition of the situation for endocrine stress responses.

Stability of the disulfide bond in an avidin-biotin linked chimeric peptide during in vivo transcytosis through brain endothelial cells.

Drug delivery of potential neuropharmaceuticals with poor intrinsic permeability through the blood-brain barrier (BBB), such as peptides, is facilitated by coupling to a vector that undergoes receptor-mediated transcytosis through the endothelial cells of brain microvessels. When cleavable disulfide linkers are used in the synthesis of such "chimeric peptides", it is crucial that the S-S-bridge is stable during transcytosis. Cleavage within endothelial cells could result in sequestration of the drug moiety instead of passage through the BBB. In the present study the metabolically stable opioid peptide [3 H]DALDA ([3 H]Tyr-DArg-Phe-Lys-NH2 ) was used as a model drug. It was monobiotinylated with the cleavable biotin reagent sulfosuccinimidyl 2-(biotinamido)ethyl-1,3'-dithiopropionate (NHS-SS-biotin) to obtain bio-[3 H]DALDA. The biotinylated peptide was then bound to a vector for brain delivery after intravenous injection in rats, a covalent conjugate of streptavidin and the transferrin receptor monoclonal antibody, OX26. Compared to peptide without vector, brain uptake of bio-[3 H]DALDA after was increased 18-fold to reach 0.12% of the injected dose per g tissue. Transcranial microdialysis was performed for 60 min after an intravenous bolus of chimeric peptide, followed by reverse phase HPLC of dialysate. Stability of the chimeric peptide during transport through the BBB into brain extracellular fluid was concluded from the absence of a peptide peak generated by disulfide cleavage.

Blood-brain barrier transport of neuropeptides: analysis with a metabolically stable dermorphin analogue.

To avoid the confounding effect of metabolic degradation, the stable mu-opioid peptide agonist [D-Arg2,Lys4]-dermorphin analogue (DALDA) was used to quantitate blood-brain barrier (BBB) permeability by intravenous injection and internal carotid artery perfusion techniques. With intravenous injection, the BBB permeability-surface area products for [3H]DALDA (0.84 +/- 0.13 microliters.min-1.g-1) and [14C]sucrose (0.39 +/- 0.05 microliters.min-1.g-1) correlated with the lipid solubility of the two molecules: the 1-octanol-Ringer partition coefficient for DALDA was approximately 2 log orders greater than that for sucrose. The brain delivery of [3H]DALDA at 30 min after intravenous administration was 0.019 +/- 0.002% of the injected dose per gram, and analgesia was induced with a 5-mg/kg dose administered systemically. In contrast to the result after intravenous injection, the BBB permeability-surface area product for DALDA estimated with the internal carotid artery perfusion technique was manyfold greater. This was due to nonspecific absorption of the peptide into the cerebral microvasculature, which precluded use of the capillary depletion technique to study transcytosis through the BBB after internal carotid artery perfusion. The present studies show that the brain delivery of a metabolically stable peptide, such as DALDA, is comparable to that for sucrose, correlates with lipid solubility, and is mediated by a nonsaturable mechanism, probably free diffusion.

Stereoselectivity of cholinesterase inhibition by galanthamine and tolerance in humans.

The effect of galanthamine (GAL) and its 2 major metabolites on human cholinesterases has been explored. Epigalanthamine, a diastereomer of GAL, was 130-times less potent in vitro in its effect on acetylcholinesterase (AChE) in erythrocytes than the parent compound, and it did not differ significantly from the ketone galanthaminone. In vivo, the maximal 36-55% inhibition of AChE was approached 30 min after oral administration of 10 mg GAL. The duration of the catalytic inhibition corresponded to an elimination half-life of approximately 5-7 h. GAL was well tolerated in 8/8 healthy volunteers, and 3/4 Alzheimer patients tolerated the drug up to a daily dose of 40 mg.

Synthesis and bioactivity of monobiotinylated DALDA: a mu-specific opioid peptide designed for targeted brain delivery.

Delivery through the blood-brain barrier of opioid peptide-based therapeutic agents may be achieved with the use of conjugation of avidin and blood-brain barrier transport vectors. However, this drug delivery strategy requires that 1) the peptide is monobiotinylated and 2) the peptide is biologically active after cleavage of a disulfide linker and peptide release from the avidin-vector conjugate. Whether these criteria may be successfully fulfilled was examined in the present studies. The highly mu receptor-specific dermorphin analog, Tyr-D-Arg-Phe-Lys-NH2 (DALDA), was selectively monobiotinylated at the epsilon-NH2 group of Lys4 with the cleavable biotin linker, sulfosuccinimidyl-2-(biotinamidoethyl) 1,3'-dithioproprionate to obtain biotinylated DALDA (bio-DALDA). The N-terminal alpha-NH2 group of the peptide was protected during biotinylation with the N-9-fluorenylmethoxycarbonyl group. Cleavage of the disulfide bridge yielded the desbiotinylated derivative, desbio-DALDA. The identity of these peptides was verified by secondary ion mass spectrometry. In receptor binding assays with 3H-Tyr-D-Ala-Gly-Phe-(N-Me)-Gly-ol, the Kis of DALDA, bio-DALDA and desbio-DALDA for mu opioid receptors were determined to be 2.3 +/- 0.4, 6.5 +/- 1.1 and 4.0 +/- 0.9 nM, respectively. Binding of bio-DALDA to avidin resulted in a Ki of 14.5 +/- 2.4 nM. The i.c.v. administration of DALDA and desbio-DALDA induced potent and long-lasting analgesia in the rat tail-flick assay. It was found that 1 microgram of DALDA was equipotent to 3 micrograms of desbio-DALDA and 20 micrograms of morphine. The analgesic effect could be blocked by naloxone pretreatment. In conclusion, these studies 1) described methods for the preparation of a biologically active monobiotinylated mu opioid receptor-specific ligand and 2) demonstrated the advantages of using cleavable biotinylation of opioid peptides because the affinity of desbio-DALDA for the receptor approximated the affinity of DALDA and had a 3- to 4-fold higher affinity than did the bio-DALDA-avidin complex.

Pharmacokinetics and saturable blood-brain barrier transport of biotin bound to a conjugate of avidin and a monoclonal antibody to the transferrin receptor.

The delivery of biotinylated therapeutics through the blood-brain barrier (BBB) may be facilitated by the use of avidin-based chimeric peptide conjugates. The latter are formed by conjugating avidin to a BBB drug delivery vector, which is a protein that undergoes receptor-mediated transcytosis through the BBB. The murine OX26 monoclonal antibody to the rat transferrin receptor undergoes receptor-mediated transport through the BBB, and previous studies have shown that a [3H]biotin/avidin-OX26 conjugate is effectively transported through the BBB. However, avidin is a cationic protein, which causes a marked increase in the systemic clearance of avidin-based conjugates from the plasma compartment. The present studies describe attempts to elevate the reduced plasma area under the curve (AUC) of [3H]biotin/avidin-OX26 by preloading or coloading with unconjugated OX26 antibody or unconjugated avidin. Both systemic clearance and BBB transport of avidin-OX26 were equally affected by OX26 preloading or coloading; this had inverse effects on the plasma AUC and the BBB permeability surface area product with no resulting change in the fractional delivery of [3H]biotin to brain. Conversely, avidin coloading preferentially reduced brain clearance of the [3H]biotin/avidin-OX26 conjugate, without substantial alteration in the plasma AUC and greatly reduced the fractional delivery of [3H]biotin to brain. In summary, these studies show that the use of avidin-based vectors results in rapid systemic clearance, which causes a reduction in the delivery of [3H]biotin to brain, despite a comparable BBB permeability coefficient for either the unconjugated OX26 antibody or the avidin-OX26 conjugate.

In vivo cleavability of a disulfide-based chimeric opioid peptide in rat brain.

Brain delivery of systemically administered neuropeptide drugs may be achieved by the synthesis of chimeric peptides, wherein the peptide is coupled to transport vectors via avidin-biotin technology. The present study focuses on factors that optimize the linkage of drugs to transport vectors. The vector is the OX26 monoclonal antibody to the transferrin receptor, and the model peptide used in these studies is [Lys7]dermorphin (K7DA). The K7DA is monobiotinylated at the epsilon-amino group of the Lys7 residue with either a cleavable linker, e.g., disulfide, using NHS-SS-biotin, or a noncleavable linker, e.g., amide, using NHS-XX-biotin. Disulfide cleavage of the biotinylated derivative yields the desbiotinylated peptide, which is thiolated. Structures of the K7DA analogues were confirmed by secondary ion mass spectrometry. The biotinylated peptides were coupled to a thiol-ether conjugate of the OX26 antibody and either neutral avidin (NLA) or streptavidin. The binding constants (Ki) of the K7DA, the biotinylated K7DA (bio-XX-K7DA), the desbiotinylated K7DA, and the bio-XX-KD7A conjugated to NLA-OX26 were 0.62 +/- 0.14, 1.59 +/- 0.27, 1.24 +/- 0.24, and > 10 nM, respectively, and were determined with a mu-opioid peptide radioreceptor assay. Comparable results were obtained with in vivo tail-flick analgesia testing following intracerebroventricular (icv) injection of opioid chimeric peptides. Reversibility of pharmacologic action of thiolated peptide was demonstrated by icv naloxone administration. The cleavability of the disulfide linker in vivo in rat plasma and brain was assessed with gel filtration HPLC and internal carotid artery perfusion of labeled opioid chimeric peptides.(ABSTRACT TRUNCATED AT 250 WORDS)

Time course of ACTH 4-10 effects on human attention.

Previous studies have demonstrated that the adrenocorticotropic hormone (ACTH) and its 4-10 fragment are potent modulators of attention in humans. In these studies the Nd component ('negative displacement') of the stimulus-evoked brain potential (event-related potential, ERP), which is a neurophysiological indicator of selective attention, proved most sensitive to effects of these peptides. The Nd decreased in amplitude linearly with logarithmically increasing doses of ACTH 4-10 applied. Here we report results complementing these previous findings by establishing a time course of the effects of ACTH 4-10 on human attention. In a double-blind cross-over study, attention performance on a dichotic listening task was tested in 18 healthy volunteers, after having received intravenously either placebo or 1 mg ACTH 4-10. Tests took place 0, 30, 90, 180, 330, 600 min, 24, 48 and 72 h after drug intake and took about 15 min. Each subject was tested on 4 of these occasions, selected according to a random schedule. During task performance ERPs to the different task stimuli were recorded. ERPs were used to determine the Nd (an indicator of controlled processing of stimuli which the subjects selectively attended to) and the mismatch negativity (MMN) reflecting automatic processing of stimulus deviance. ACTH 4-10 diminished the Nd significantly at 30 min after treatment administration. On none of the other occasions significant changes in Nd occurred. MMN was not affected by the peptide. The results indicate a short-term action of ACTH 4-10 on selective attention, which takes about 30 min to develop and does not last longer than 90 min after treatment.

The behaviorally active peptide ACTH 4-10: measurement in plasma and pharmacokinetics in man.

A specific radioimmunoassay for the quantitative measurement of ACTH 4-10 and a procedure for its extraction from plasma have been developed. Its pharmacokinetics was studied in eight healthy male volunteers given ACTH 4-10 125 micrograms/kg body weight as a bolus i.v. injection, by infusion and intranasally. Following the i.v. bolus, plasma levels rapidly declined biexponentially, with half-lives of 0.39 +/- 0.05 min for the alpha-phase and 3.84 +/- 1.5 min for the beta-phase (mean +/- SD). The constant rate i.v. infusion yielded steady-state levels between 0.74 and 5.06 ng/ml plasma. Administered as intranasal spray, absorption of intact ACTH 4-10 was low and variable (maximal bioavailability 7.6%). The results are discussed in relation to the dose-dependent effects of ACTH 4-10 on the auditory evoked potential.