Stimuli-responsive graphene-based hydrogel driven by disruption of triazine hydrophobic interactions.

The study reported here concerns the preparation of a novel graphene-diaminotriazine (G-DAT) nanocomposite hydrogel for application in the drug delivery field. The hybrid nature of this material is founded on two key elements: the presence of the DAT backbone induced the formation of hydrophobic regions that allowed efficient loading of a series of drugs of increasing hydrophobicity (Metronidazole, Benzocaine, Ibuprofen, Naproxen and Imipramine), while simultaneously endowing swelling-induced pH-responsiveness to the hydrogel. Additionally, the incorporation of graphene was found to interfere with these hydrophobic domains through favourable non-covalent interactions, thus leading to the partial disruption of these aggregates. As a consequence, graphene facilitated and enhanced the release of model hydrophobic drug Imipramine in a synergistic manner with the pH trigger, and increased the swelling capacities and improved mechanical performance. This hybrid hydrogel can therefore be envisaged as a proof-of-concept system for the release of hydrophobic compounds in the field of drug delivery.

DPCPX, a selective adenosine A1 receptor antagonist, enhances the antidepressant-like effects of imipramine, escitalopram, and reboxetine in mice behavioral tests.

The main goal of the present study was to evaluate the influence of the adenosine A1 receptor (A1R) antagonist - DPCPX - on depressive-like behavior in mice, as well as the effect of DPCPX on the activity of imipramine, escitalopram, and reboxetine, each at non-effective doses. The influence of DPCPX on behavior and its influence on the activity of selected antidepressants was evaluated in the forced swim test (FST) and the tail suspension test (TST) in mice. Locomotor activity was measured to verify and exclude false-positive data obtained in the FST and TST. Moreover, serum and brain concentrations of tested antidepressants were determined using HPLC. DPCPX, at doses of 2 and 4 mg/kg, exhibited antidepressant activity in the FST and TST, which was not related to changes in the spontaneous locomotor activity. Co-administration of DPCPX with imipramine, escitalopram, or reboxetine, each at non-active doses, significantly reduced the immobilization period in the FST and TST in mice, which was not due to the increase in locomotor activity. Both antagonists of 5-HT receptors (WAY 100635 and ritanserin) completely antagonized the effect elicited by DPCPX in the behavioral tests. Results of assessment of the nature of the interaction between DPCPX and test drugs show that in the case of DPCPX and imipramine or reboxetine, there were pharmacodynamic interactions, whereas the DPCPX-escitalopram interaction is at least partially pharmacokinetic in nature. Presented outcomes indicate that an inhibition of A1Rs and an increase of monoaminergic transduction in the CNS may offer a novel strategy for the development of antidepressant drugs.

Investigation on the Interactions between Various Drugs and Aojiru (Green Juice) Using a Simple Centrifugation Method.

The objective of this study was to evaluate the interactions between various drugs and aojiru (green juice), a popular health food in Japan, using a simple centrifugation method. The mixture of drug solution and aojiru suspension was gently shaken and centrifuged. The drug concentration in the supernatant fluid was then determined by HPLC. The concentration of rhodamine 123 (Rho-123), a model compound, in the supernatant fluid significantly decreased after mixing with aojiru, indicating extensive binding of Rho-123 to the insoluble components of aojiru. When administered into rat small intestinal loops together with aojiru, the plasma Rho-123 concentrations became much smaller than those when administered alone. This result strongly suggested that a strong interaction observed in vitro was well reflected in modulated absorption. Among seven drugs tested, chlorpromazine and imipramine exerted binding properties to aojiru similar to or greater than Rho-123. As a small part of both Rho-123 and imipramine was released when the aojiru precipitate was resuspended, their binding to aojiru was considered to be tight. The binding of diltiazem, fexofenadine, glibenclamide, metformin, and norfloxacin to aojiru was much weaker or almost negligible compared with that of chlorpromazine and imipramine. The present results suggest that aojiru can decrease the intestinal absorption of some clinically relevant drugs through tight binding in the small intestine and that the present centrifugation method is useful for predicting in vivo interactions between drugs and aojiru.

Hepatic Flavin-Containing Monooxygenase 3 Enzyme Suppressed by Type 1 Allergy-Produced Nitric Oxide.

Flavin-containing monooxygenases (FMOs) are major mammalian non-cytochrome P450 oxidative enzymes. T helper 2 cell-activated allergic diseases produce excess levels of nitric oxide (NO) that modify the functions of proteins. However, it remains unclear whether allergy-induced NO affects the pharmacokinetics of drugs metabolized by FMOs. This study investigated alterations of hepatic microsomal FMO1 and FMO3 activities in type 1 allergic mice and further examined the interaction of FMO1 and FMO3 with allergy-induced NO. Imipramine (IMP; FMO1 substrate) N-oxidation activity was not altered in allergic mice with high serum NO and immunoglobulin E levels. At 7 days after primary sensitization (PS7) or secondary sensitization (SS7), benzydamine (BDZ; FMO1 and FMO3 substrate) N-oxygenation was significantly decreased to 70% of individual controls. The expression levels of FMO1 and FMO3 proteins were not significantly changed in the sensitized mice. Hepatic inducible NO synthase (iNOS) mRNA level increased 5-fold and 15-fold in PS7 and SS7 mice, respectively, and hepatic tumor necrosis factor-α levels were greatly enhanced. When a selective iNOS inhibitor was injected into allergic mice, serum NO levels and BDZ N-oxygenation activity returned to control levels. NO directly suppressed BDZ N-oxygenation, which was probably related to FMO3-dependent metabolism in comparison with IMP N-oxidation. In hepatic microsomes from PS7 and SS7 mice, the suppression of BDZ N-oxygenation was restored by ascorbate. Therefore, type 1 allergic mice had differentially suppressed FMO3-dependent BDZ N-oxygenation. The suppression of FMO3 metabolism related to reversible S-nitrosyl modifications of iNOS-derived NO. NO is expected to alter FMO3-metabolic capacity-limited drug pharmacokinetics in humans.

High performance liquid chromatographic determination of ultra traces of two tricyclic antidepressant drugs imipramine and trimipramine in urine samples after their dispersive liquid-liquid microextraction coupled with response surface optimization.

Dispersive liquid-liquid microextraction (DLLME) coupled with high performance liquid chromatography by ultraviolet detection (HPLC-UV) as a fast and inexpensive technique was applied to the determination of imipramine and trimipramine in urine samples. Response surface methodology (RSM) was used for multivariate optimization of the effects of seven different parameters influencing the extraction efficiency of the proposed method. Under optimized experimental conditions, the enrichment factors and extraction recoveries were between 161.7-186.7 and 97-112%, respectively. The linear range and limit of detection for both analytes found to be 5-100ng mL(-1) and 0.6ng mL(-1), respectively. The relative standard deviations for 5ng mL(-1) of the drugs in urine samples were in the range of 5.1-6.1 (n=5). The developed method was successfully applied to real urine sample analyses.

Lysosomal sequestration (trapping) of lipophilic amine (cationic amphiphilic) drugs in immortalized human hepatocytes (Fa2N-4 cells).

Lipophilic (logP > 1) and amphiphilic drugs (also known as cationic amphiphilic drugs) with ionizable amines (pKa > 6) can accumulate in lysosomes, a process known as lysosomal trapping. This process contributes to presystemic extraction by lysosome-rich organs (such as liver and lung), which, together with the binding of lipophilic amines to phospholipids, contributes to the large volume of distribution characteristic of numerous cardiovascular and central nervous system drugs. Accumulation of lipophilic amines in lysosomes has been implicated as a cause of phospholipidosis. Furthermore, elevated levels of lipophilic amines in lysosomes can lead to high organ-to-blood ratios of drugs that can be mistaken for active drug transport. In the present study, we describe an in vitro fluorescence-based method (using the lysosome-specific probe LysoTracker Red) to identify lysosomotropic agents in immortalized hepatocytes (Fa2N-4 cells). A diverse set of compounds with various physicochemical properties were tested, such as acids, bases, and zwitterions. In addition, the partitioning of the nonlysosomotropic atorvastatin (an anion) and the lysosomotropics propranolol and imipramine (cations) were quantified in Fa2N-4 cells in the presence or absence of various lysosomotropic or nonlysosomotropic agents and inhibitors of lysosomal sequestration (NH4Cl, nigericin, and monensin). Cellular partitioning of propranolol and imipramine was markedly reduced (by at least 40%) by NH4Cl, nigericin, or monensin. Lysosomotropic drugs also inhibited the partitioning of propranolol by at least 50%, with imipramine partitioning affected to a lesser degree. This study demonstrates the usefulness of immortalized hepatocytes (Fa2N-4 cells) for determining the lysosomal sequestration of lipophilic amines.

The liver partition coefficient-corrected inhibitory quotient and the pharmacokinetic-pharmacodynamic relationship of directly acting anti-hepatitis C virus agents in humans.

Pharmacokinetic-pharmacodynamic (PK-PD) data analyses from early hepatitis C virus (HCV) clinical trials failed to show a good correlation between the plasma inhibitory quotient (IQ) and antiviral activity of different classes of directly acting antiviral agents (DAAs). The present study explored whether use of the liver partition coefficient-corrected IQ (LCIQ) could improve the PK-PD relationship. Animal liver partition coefficients (Kp(liver)) were calculated from liver to plasma exposure ratios. In vitro hepatocyte partition coefficients (Kp(hep)) were determined by the ratio of cellular to medium drug concentrations. Human Kp(liver) was predicted using an in vitro-in vivo proportionality method: the species-averaged animal Kp(liver) multiplied by the ratio of human Kp(hep) over those in animals. LCIQ was calculated using the IQ multiplied by the predicted human Kp(liver). Our results demonstrated that the in vitro-in vivo proportionality approach provided the best human Kp(liver) prediction, with prediction errors of <45% for all 5 benchmark drugs evaluated (doxorubicin, verapamil, digoxin, quinidine, and imipramine). Plasma IQ values correlated poorly (r(2) of 0.48) with maximum viral load reduction and led to a corresponding 50% effective dose (ED(50)) IQ of 42, with a 95% confidence interval (CI) of 0.1 to 148534. In contrast, the LCIQ-maximum VLR relationship fit into a typical sigmoidal curve with an r(2) value of 0.95 and an ED(50) LCIQ of 121, with a 95% CI of 83 to 177. The present study provides a novel human Kp(liver) prediction model, and the LCIQ correlated well with the viral load reductions observed in short-term HCV monotherapy of different DAAs and provides a valuable tool to guide HCV drug discovery.

Anti-invasive adjuvant therapy with imipramine blue enhances chemotherapeutic efficacy against glioma.

The invasive nature of glioblastoma (GBM) represents a major clinical challenge contributing to poor outcomes. Invasion of GBM into healthy tissue restricts chemotherapeutic access and complicates surgical resection. Here, we test the hypothesis that an effective anti-invasive agent can "contain" GBM and increase the efficacy of chemotherapy. We report a new anti-invasive small molecule, Imipramine Blue (IB), which inhibits invasion of glioma in vitro when tested against several models. IB inhibits NADPH (reduced form of nicotinamide adenine dinucleotide phosphate) oxidase-mediated reactive oxygen species generation and alters expression of actin regulatory elements. In vivo, liposomal IB (nano-IB) halts invasion of glioma, leading to a more compact tumor in an aggressively invasive RT2 syngeneic astrocytoma rodent model. When nano-IB therapy was followed by liposomal doxorubicin (nano-DXR) chemotherapy, the combination therapy prolonged survival compared to nano-IB or nano-DXR alone. Our data demonstrate that nano-IB-mediated containment of diffuse glioma enhanced the efficacy of nano-DXR chemotherapy, demonstrating the promise of an anti-invasive compound as an adjuvant treatment for glioma.

Inhibition of P-glycoprotein enhances transport of imipramine across the blood-brain barrier: microdialysis studies in conscious freely moving rats.

BACKGROUND AND PURPOSE: Recent studies indicate that efflux of antidepressants by the multidrug resistance transporter P-glycoprotein (P-gp) at the blood-brain barrier (BBB) may contribute to treatment-resistant depression (TRD) by limiting intracerebral antidepressant concentrations. In addition, clinical experience shows that adjunctive treatment with the P-gp inhibitor verapamil may improve the clinical outcome in TRD. Therefore, the present study aimed to investigate the effect of P-gp inhibition on the transport of the tricyclic antidepressant imipramine and its active metabolite desipramine across the BBB. EXPERIMENTAL APPROACH: Intracerebral microdialysis in rats was used to monitor brain levels of imipramine and desipramine following i.v. imipramine administration, with or without pretreatment with one of the P-gp inhibitors verapamil or cyclosporin A (CsA). Plasma drug levels were also determined at regular intervals. KEY RESULTS: Pretreatment with either verapamil or CsA resulted in significant increases in imipramine concentrations in the microdialysis samples, without altering imipramine plasma pharmacokinetics. Furthermore, pretreatment with verapamil, but not CsA, led to a significant elevation in plasma and brain levels of desipramine. CONCLUSIONS AND IMPLICATIONS: The present study demonstrated that P-gp inhibition enhanced the intracerebral concentration of imipramine , thus supporting the hypothesis that P-gp activity restricts brain levels of certain antidepressants, including imipramine. These findings may help to explain reports of a beneficial response to adjunctive therapy with verapamil in TRD.

Neuron density and serotonin receptor binding in prefrontal cortex in suicide.

Although serotonin receptor and cytoarchitectonic alterations are reported in prefrontal cortex (PFC) in suicide and depression, no study has considered binding relative to neuron density. Therefore, we measured neuron density and serotonin transporter (SERT), 5-HT1A and 5-HT2A binding in matched suicides and controls. Suicides and normal controls (n=15 matched pairs) were psychiatrically characterized. Neuron density and binding were determined in dorsal [Brodmann area (BA) 9] and ventral (BA 47) PFC by stereology and quantitative autoradiography in near-adjacent sections. Binding index was defined as the ratio of receptor binding to neuron density. Suicides had lower neuron density in the gyrus of both areas. The binding index was lower for SERT in BA 47 but not in BA9; the 5-HT1A binding index was higher in BA 9 but not in BA 47, while the 5-HT2A binding index was not different between groups. SERT binding was lower in suicides in BA 47 but not BA 9, while 5-HT1A binding was higher in BA 9 but not BA 47. SERT binding negatively correlated with 5-HT1A binding in BA 47 in suicides. Neuron density decreased with age. The 5-HT1A binding index was higher in females than males. We found lower neuron density and lower SERT binding index in both PFC regions in suicides. More 5-HT1A binding with less SERT binding and the negative correlation in depressed suicides suggests post-synaptic receptor up-regulation, and it is independent of the difference in neuron density. Thus, abnormalities in both cortical neurons and in their serotonergic innervation are present in suicides and future studies will need to determine whether cortical changes reflect the trophic effect of altered serotonin innervation.

Serotonergic and noradrenergic pathways are required for the anxiolytic-like and antidepressant-like behavioral effects of repeated vagal nerve stimulation in rats.

BACKGROUND: Vagal nerve stimulation (VNS) is used for treatment-refractory depression, but there are few preclinical studies of its effects when administered repeatedly over time using clinically relevant stimulation parameters in nonanesthetized animals. METHODS: The novelty-suppressed feeding test (NSFT) and forced swim test (FST) were used to evaluate the anxiolytic- and antidepressant-like potential of VNS in rats, respectively. The behavioral effects of VNS were compared with those of desipramine (DMI; 10 mg/kg/day) and sertraline (7.5 mg/kg/day) administered via osmotic minipump. Such experiments were carried out in intact rats as well as those that had selective destruction of either serotonin or noradrenergic neurons in brain caused by the neurotoxins, 5,7-dihyroxytryptamine (5,7-DHT), or 6-hydroxydopamine (6-OHDA). RESULTS: Repeated administration of VNS, DMI, and sertraline decreased latency to feed in the NSFT. In the FST, repeated VNS, DMI, and sertraline caused decreased immobility; the VNS-induced decrease in immobility resulted from increases in both swimming and climbing behaviors. Effects of VNS and sertraline, but not DMI, in both the NSFT and the FST were abolished in rats treated with 5,7-DHT. Effects of DMI in both behavioral tests, but not those of sertraline, were abolished in 6-OHDA treated rats. VNS effects on immobility and climbing in the FST were not blocked in the 6-OHDA-treated rats. There was no significant difference in locomotor activity caused by any of the treatments or by the lesions. CONCLUSIONS: Serotonergic nerves are required for repeated VNS-induced anxiolytic- and antidepressant-like effects. Noradrenergic nerves can also be activated by VNS to cause its anxiolytic-like effect.

Development of a novel high-throughput analytical methodology, multiple injection method, for quantitative analysis in drug metabolism and pharmacokinetic studies using liquid chromatography with tandem mass spectrometry.

In this study, we developed a novel methodology, multiple injection method (MIM), for higher-throughput screening of compounds by liquid chromatography-tandem mass spectrometry (LC-MS/MS). MIM involves continuous injections of multiple samples containing a different compound respectively into the column, and then temporarily trapping of analytes at the column head in high-pressure liquid chromatography (HPLC) system. This is followed by elution of all the compounds from the column and detection of them by MS/MS. In this study, fexofenadine, verapamil, risperidone, ondansetron, and imipramine were used as model compounds to investigate the effectiveness of MIM in pharmacokinetic studies. Analytical time of validation samples of these model compounds could be shortened to one third by MIM, compared with the conventional method. In addition, both the accuracy and precision of MIM met the general criteria for quantitative analysis. The peak intensity was found to be unaffected by overlapping compounds even if they have wide range of ionization efficiency. As a result of the comparison of MIM and conventional method in the analysis of samples in pharmacokinetic studies using model compounds, no difference was shown in the quantification values. Consequently, this method has some advantages, reduction of analytical time, the improvement of sensitivity, and the simplicity of system, compared to the conventional methods. MIM should be very useful and powerful method for drug development without an additional hardware and can be used for the measurement of compounds in biological samples for pharmacokinetic studies, especially it greatly contributes to accelerating drug development in its discovery stages.

The tricyclic antidepressants amitriptyline, nortriptyline and imipramine are weak antagonists of human and rat alpha1B-adrenoceptors.

Although it is long known that the tricyclic antidepressants amitriptyline, nortriptyline and imipramine inhibit the noradrenaline transporter and alpha(1)-adrenoceptors with similar affinities, which may lead to self-cancelling actions, the selectivity of these drugs for alpha(1)-adrenoceptor subtypes is unknown. The present study investigates the selectivity of amitriptyline, nortriptyline and imipramine for human recombinant and rat native alpha(1)-adrenoceptor subtypes. The selectivity of amitriptyline, nortriptyline and imipramine was investigated in HEK-293 cells expressing each of the human alpha(1)-subtypes and in rat native receptors from the vas deferens (alpha(1A)), spleen (alpha(1B)) and aorta (alpha(1D)) through [(3)H]prazosin binding, and noradrenaline-induced intracellular Ca(2+) increases and contraction assays. Amitriptyline, nortriptyline and imipramine showed considerably higher affinities for alpha(1A)- (approximately 25- to 80-fold) and alpha(1D)-adrenoceptors (approximately 10- to 25-fold) than for alpha(1B)-adrenoceptors in both contraction and [(3)H]prazosin binding assays with rat native and human receptors, respectively. In addition, amitriptyline, nortriptyline and imipramine were substantially more potent in the inhibition of noradrenaline-induced intracellular Ca(2+) increases in HEK-293 cells expressing alpha(1A)- or a truncated version of alpha(1D)-adrenoceptors which traffics more efficiently towards the cell membrane than in cells expressing alpha(1B)-adrenoceptors. Amitriptyline, nortriptyline and imipramine are much weaker antagonists of rat and human alpha(1B)-adrenoceptors than of alpha(1A)- and alpha(1D)-adrenoceptors. The differential affinities for these receptors indicate that the alpha(1)-adrenoceptor subtype which activation is most increased by the augmented noradrenaline availability resultant from the blockade of neuronal reuptake is the alpha(1B)-adrenoceptor. This may be important for the behavioural effects of these drugs.

Selective alterations of the CB1 receptors and the fatty acid amide hydrolase in the ventral striatum of alcoholics and suicides.

Recent studies in rodents have suggested a role for the central endocannabinoid system in the regulation of mood and alcohol related behaviors. Alcohol use disorder is often associated with suicidal behavior. In the present study, we examined whether abnormalities in the endocannabinoid system in the ventral striatum are associated with alcohol dependence and suicide. The levels of CB1 receptors, receptor-mediated G-protein signaling, and activity and level of the fatty acid amide hydrolase (FAAH) were analyzed postmortem in the ventral striatum of alcohol-dependent nonsuicides (CA, n=9), alcohol-dependent suicides (AS, n=9) and nonpsychiatric controls (C, n=9). All subjects underwent a psychological autopsy, and toxicological and neuropathological examinations. The levels of the CB1 receptors and the CB1 receptor-mediated G-protein signaling were significantly lower in the ventral striatum of CA compared to the control group. However, these parameters were elevated in AS when compared to CA group. The activity of FAAH enzyme was lower in CA compared to the control group while it was found to be significantly higher in AS compared with CA group. These findings suggest that alcohol dependence is associated with the downregulation of the CB1 receptors, while suicide is linked to the upregulation of these receptors in the ventral striatum. Alteration in the activity of FAAH enzyme that regulates the anandamide (AEA) content might in turn explain differences in the CB1 receptor function in alcohol dependence and suicide. These findings may have etiological and therapeutic implications for the treatment of alcohol addiction and suicidal behavior.

Comparison of fused-core and conventional particle size columns by LC-MS/MS and UV: application to pharmacokinetic study.

The chromatographic performance of fused-core (superficially porous) HPLC packing materials was compared with conventional fully porous particle materials for LC-MS/MS analysis of two pharmaceuticals in rat plasma. Two commercially available antidepressants, imipramine and desipramine, were assayed using a conventional analytical C(18) column (5 microm, 2.0 mm x 30 mm) and a fused-core C(18) column (2.7 microm, 2.1 mm x 30 mm). Retention time, column efficiency, pressure drop, resolution, and loading capacity were compared under the same operating conditions. The fused-core column demonstrated reduced assay time by 34% and 2-3-fold increased efficiency (N). Loading capacity up to 25 microl of extract injected on column showed no peak distortion. The registered back-pressure from a flow rate of 1.0 ml/min did not exceed 3400 psi making it compatible with standard HPLC equipment (typically rated to 6000 psi). Two mobile phases were examined, and morpholine as an organic base modifier yielded a 2-5-fold increase in S/N near the limit of detection over triethylamine. The 2.7 microm fused-core column was applied to the analysis of imipramine and desipramine in extracted, protein precipitated rat plasma by LC-MS/MS. The calibration curves were linear in the concentration range of 0.5-1000 ng/ml for both imipramine and desipramine. Intra-run precisions (%CV) and accuracies (%bias) were within +/-7.8% and +/-7.3% at three QC levels and within 14.7% and 14.4% at the LOQ level for both analytes. Following a single method qualification run, the method was applied to the quantitation of pharmacokinetic study samples after oral administration of imipramine to male rats.

Verapamil in treatment resistant depression: a role for the P-glycoprotein transporter?

INTRODUCTION: Verapamil is a calcium channel blocker that also inhibits the P-glycoprotein (Pgp) membrane transporter. We have found that administration of verapamil with a recognised antidepressant improves clinical outcome in previously treatment resistant cases despite the fact that verapamil does not possess inherent antidepressant activity. In this study we examined the hypothesis that the antidepressant-like effects of verapamil are mediated through its blockade of the Pgp transporter in the blood brain barrier (BBB). METHODS: Following pre-treatment with verapamil (20 mg/kg) or a saline solution male Sprague Dawley rats were injected with imipramine (15 mg/kg). Two hours later, the animals were sacrificed, trunk blood collected and brain regions dissected out. High performance liquid chromatography (HPLC) was used to quantitate antidepressant drug concentrations in all samples. RESULTS: Verapamil pre-treatment significantly elevated imipramine concentrations in all brain regions studied. The effect was most pronounced in the brainstem and frontal cortex where we observed in excess of a doubling in the brain region: serum ratios. CONCLUSION: Our results verify inhibition of Pgp as a potential mechanism of action for verapamil during treatment resistant depression. The implications of these findings are discussed in the context of novel treatment strategies in depression.

Alteration of skin hydration and its barrier function by vehicle and permeation enhancers: a study using TGA, FTIR, TEWL and drug permeation as markers.

Vehicles and permeation enhancers (PEs) used in transdermal drug delivery (TDD) of a drug can affect skin hydration, integrity and permeation of the solute administered. This investigation was designed to study the effect of the most commonly used vehicles and PEs on rat skin hydration, barrier function and permeation of an amphiphilic drug, imipramine hydrochloride (IMH). An array of well-established techniques were used to confirm the findings of the study. Thermogravimetric analysis (TGA) and Fourier transform infrared (FTIR) spectroscopy were used to determine changes in skin hydration. Alteration of the stratum corneum (SC) structure was investigated using FTIR studies. To monitor the barrier function alteration, transepidermal water loss (TEWL) measurement and permeation studies were performed. Our findings indicate that with hydration, there was an increase in the bound water content of the skin, and pseudoequilibrium of hydration (a drastic decrease in hydration rate) was achieved at around 12 h. Hydration increased the ratio between amide-I and amide-II peaks in FTIR and reduced the C-H stretching peak area. Both propylene glycol (PG) and ethanol (EtOH) dehydrated skin, with the latter showing a predominant effect. Furthermore, it was confirmed that PG and EtOH decreased the bound water content due to alteration in the protein domains and extraction of SC lipids, respectively. The effect of hydration on the SC was found to be similar to that reported for temperature. Permeation studies revealed that the dehydration caused by vehicles decreased IMH flux, whereas the flux was enhanced by PEs. The role of partition was predominant for the permeation of IMH through dehydrated skin. A synergistic effect was observed for PG and menthol in the enhancement of IMH. Further findings provided strong evidence that PG affects protein domains and EtOH extracts lipids from the bilayer. Both PG and EtOH, with or without PEs, increased TEWL. Initial TEWL was well correlated with the flux of IMH through the same skin. It was found that both PG and EtOH affect the permeation of solute and TEWL by dehydration. The experiments also proved that the initial TEWL value has a strong potential as a predictive tool for the permeation of the solute.

Influence of serotonin 5-HT(7) receptor blockade on the behavioral and neurochemical effects of imipramine in rats.

The aim of the present study was to examine the effect of the selective 5-HT(7) receptor antagonist (2R)-1-[(3-hydroxyphenyl)sulfonyl]-2-[2-(4-methyl-1-piperidinyl)ethyl]pyrrolidine (SB-269970), administered alone or in combination with imipramine, on the immobility time of rats in the forced swim test as well as on the extracellular levels of dopamine (DA), noradrenaline (NA), serotonin (5-HT) and their metabolites in the prefrontal cortex of freely moving rats. Both compounds were administered intraperitoneally (ip). Like imipramine (30 mg/kg, but not 20 mg/kg), SB-269970 (1.25 and 2.5 mg/kg, but not 0.625 mg/kg) significantly shortened the immobility time of rats without affecting their exploratory locomotor activity measured in the open field test. SB-269970 (0.625 and 1.25 mg/kg) raised the extracellular levels of DA, NA, 5-HT and their metabolites in rat prefrontal cortex. In that structure, imipramine (20 mg/kg) produced an increase in all the neurotransmitters measured, but failed to affect the levels of their metabolites. A combination of the inactive doses of SB-269970 (0.625 mg/kg) and imipramine (20 mg/kg) found in the forced swim test produced antidepressant-like effect, which did not stem from the increased exploratory locomotor activity. At the same time, that combination voked a vast increase in the output of NA - but not DA and 5-HT - compared to the effects of both those drugs given alone. These results open up a possibility that the stimulating effect of SB-269970 on DA, NA and 5-HT transmission in the prefrontal cortex plays some role in the antidepressant-like activity of this compound. Moreover, these findings suggest that the increase in cortical NA level seems to account for the anti-immobility action observed after joint administration of the selective 5-HT(7) receptor antagonist and imipramine in rats.

Pharmacokinetic and pharmacodynamic studies of drug interaction following oral administration of imipramine and sodium alginate in rats.

Recently, the use of health foods has increased due to growing interest in health maintenance. Previous in vitro studies have shown some drugs to be adsorbed by sodium alginate, a dietary fiber, and that such adsorption was marked with tricyclic antidepressants, such as imipramine. This study investigated the pharmacokinetic and pharmacological interactions between imipramine and sodium alginate in rats. The simultaneous administration of imipramine (30 mg/kg, oral (p.o.)) and sodium alginate (3.0%, p.o.) decreased the antidepressant-like activity of imipramine in a forced swimming test. In the rats administrated imipramine and 0.3%, 1.0%, or 3.0% sodium alginate, the geometric mean ratio of the Cmax values of imipramine was 72% [90% confidence intervals (CI) = 53-91%], 64% (90% CI = 47-80%), and 58% (90% CI = 50-67%), respectively. The geometric mean ratio of the AUC(0-6) values of imipramine were 68% (90% CI = 56-80%), 74% (90% CI = 60-89%), and 87% (90% CI = 73-102%), respectively. The decrease in Cmax and AUC(0-6) was judged to be significant with a 90% CI outside the 80-125% boundaries. In addition, the Tmax value of imipramine significantly increased (P < 0.05) by coadministration with 3.0% sodium alginate. These results suggested that simultaneous administration of sodium alginate decreased the serum concentration and pharmacological action of imipramine, through a delay in its absorption. Although the clinical relevance of these findings is unclear, it is important to pay considerable attention to the interactions between imipramine and sodium alginate.

New method for the simultaneous estimation of intrinsic hepatic clearance and protein binding by matrix inhibition.

The purpose of this study was to develop a method for estimating the hepatic clearance (CL(h)) without using a protein binding test. This method allows the simultaneous evaluation of the intrinsic hepatic clearance (CL(int)) with a correction for microsomal binding, and the free fraction in the serum (fu). It uses the decrease in metabolic velocity achieved by decreasing the free fraction of a compound in the incubation mixture (fu(inc)) by the addition of serum, and by changing the microsomal protein concentration. This method is denoted as the 'matrix inhibition method', because it uses the inhibition of the metabolic velocity by the incubation matrix. The metabolic rates of eight compounds (diazepam, imipramine, warfarin, and compounds A-E) were evaluated under several incubation conditions using rat serum and microsomes. The correlation of CL(int) evaluated using the method and using equilibrium dialysis after the CL(int) was corrected for microsomal binding was r = 0.968. The correlation of fu . CL(int) was r = 0.996. Although the method required a high enough fu and fu(microsomes) difference among the reaction conditions for each compound, it could evaluate CL(int) and fu simultaneously and easily by adding additional reaction conditions to the metabolic stability tests performed in ADME screening.