Brain Microdialysis Method Research Articles

Pharmacokinetic comparative study of GAS with different concentration of tetramethylpyrazine and ferulic acid on liver–yang hyperactivity migraine model by blood–brain microdialysis method

Gastrodia elata and Ligusticum chuanxiong are used to treat primary headaches for many years. Gastrodin (GAS) and tetramethylpyrazine (TMP)/ferulic acid (FA) are the main active ingredients of Gastrodia elata and Ligusticum chuanxiong, respectively. Previous studies demonstrated the pharmacokinetics of GAS, TMP, and FA in the blood and brain interstitial fluids (BIF) in healthy animals but not in animal model with liver–yang hyperactivity migraine. Hence, this study examined the pharmacokinetics of GAS after its oral administration in the presence of different concentrations of TMP and FA in animals with liver–yang migraine hyperactivity. In the control group, GAS was administrated without TMP and FA. Pharmacokinetic parameters were determined using the blood-brain microdialysis in combination with the high-performance liquid chromatography method. Results revealed that the maximum drug concentrations (Cmax) in the serum, area under curve (AUC), and mean residence time (MRT) of GAS decreased in normal animals, whereas Cmax increased significantly in model animals. These findings indicate that varying concentrations of TMP and FA play an important role in the pharmacokinetics of GAS in both normal and migraine model animals, validating the utility of the ancient formulation.

Pharmacokinetic comparative study of tetramethylpyrazine and ferulic acid and their compatibility with different concentration of gastrodin and gastrodigenin on blood–stasis migraine model by blood–brain microdialysis method

Tianma pills, a traditional formula made from Ligusticum chuanxiong and Gastrodia elata, are efficacious for the treatment of primary headache. Tetramethylpyrazine (TMP) and Ferulic acid (FA) are the bioactive ingredients of Ligusticum chuanxiong, while Gastrodin and Gastrodigenin are the bioactive ingredients of Gastrodia elata. Pharmacokinetic assessment of TMP, FA, gastrodin or gastrodigenin in blood or brain interstitial fluid (BIF) has been reported in healthy animals. However, the pharmacokinetic properties of TMP and FA have not been studied when they are co–administered in a blood–stasis migraine model. The present research investigated the pharmacokinetic behavior of TMP and FA after oral administration in the presence of different concentrations of gastrodin and gastrodigenin in a blood–stasis migraine model. Pharmacokinetic parameters were determined using blood–brain microdialysis in combination with the UHPLC–MS method. Compared to the control group, in which TMP and FA were administrated without gastrodin or gastrodigenin, the T1/2, MRT, Cmax and AUC0–∞ of TMP and FA were increased. These results indicate that varying concentrations of gastrodin and gastrodigenin play an important role in affecting the pharmacokinetics of TMP and FA. Low concentrations of gastrodin and gastrodigenin (similar to those found in Tianma pills) were more efficacious, validating the utility of the ancient formulation.

Brain Microdialysis Study of Vancomycin in the Cerebrospinal Fluid After Intracerebroventricular Administration in Mice

Vancomycin (VCM) is an important antibiotic for treating methicillin-resistant Staphylococcus aureus (MRSA) infections. To treat bacterial meningitis caused by MRSA, it is necessary to deliver VCM into the meninges, but the rate of VCM translocation through the blood-brain barrier is poor. Additionally, high doses of intravascularly (i.v.) administered VCM may cause renal impairments. Thus, VCM is sometimes administered intracerebroventricularly (i.c.v.) for clinical treatment. However, information on the VCM pharmacokinetics in cerebrospinal fluid (CSF) after i.c.v. administration is lacking. In the present study, we evaluated the VCM pharmacokinetics in the CSF and systemic circulation after i.c.v. compared to that after i.v. administration, using the brain microdialysis method in mice. VCM administered via i.c.v. showed a highly selective distribution in the CSF, without migration to systemic circulation. Moreover, to assess renal impairments after i.c.v. administration of VCM, we histologically evaluated damage to the mouse kidney by hematoxylin and eosin staining. No significant morphological change in the kidney was observed in the i.c.v. administration group compared to that in the i.v. administration group. Our results demonstrate that i.c.v. administration of VCM can be partially prevented from entering the systemic circulation to prevent renal impairments caused by VCM.

Use of Subcutaneous and Intraperitoneal Administration Methods to Facilitate Cassette Dosing in Microdialysis Studies in Rats.

Microdialysis is a powerful technique allowing for real-time measurement of unbound drug concentrations in brain interstitial fluid in conscious animals. Use of microdialysis in drug discovery is limited by high resource requirement and low throughput, but this may be improved by cassette dosing. Administering multiple compounds intravenously of diverse physiochemical properties, it is often very challenging and time consuming to identify a vehicle that can dissolve all of the compounds. To overcome this limitation, the present study explores the possibility of administering a cassette dose of nine diverse compounds (carbamazepine, citalopram, desmethylclozapine, diphenhydramine, gabapentin, metoclopramide, naltrexone, quinidine, and risperidone) in suspension, rather than in solution, by intraperitoneal and subcutaneous routes, and determining if this is a viable option for assessing blood-brain barrier penetration in microdialysis studies. Repeated hourly subcutaneous dosing during the 6-hour microdialysis study allowed for the best attainment of distributional equilibrium between brain and plasma, resulting in less than a 2-fold difference in the unbound brain to unbound plasma concentration ratio for the cassette dosing method versus discrete dosing. Both subcutaneous and intraperitoneal repeated dosing can provide a more practical substitute for intravenous dosing in determining brain penetration of a cassette of diverse compounds in brain microdialysis studies. The results from the present study demonstrate that dosing compounds in suspension represents a practical approach to eliminating the technical challenge and labor-intensive step of preparation of solutions of a mixture of compounds and will enable the use of the cassette brain microdialysis method in a central nervous system drug discovery setting.

Comparative brain pharmacokinetic study of Jiaotai pills in normal and insomnic rats using brain microdialysis combinated with LC–MS/MS

ObjectiveTo compare the brain pharmacokinetics of five protoberberine-type alkaloids (i.e. berberine, palmatine, coptisine, epiberberine, and jatrorrhizine), which were the main bioactive constituents of Jiaotai Pills (JTP), in normal and insomnic rats orally administrated with JTP. MethodsThe detection was conducted by a fully validated liquid chromatography-tandem mass spectrometry combinated with brain microdialysis method. Brain microdialysis probes were inserted into the hippocampus of rats. JTP extracts were administrated intragastrically and then brain microdialysates were collected at 30 min time intervals for 10 h. The separation of the five protoberberine-type alkaloids was carried out on a BDS Hypersil C18 column using a mobile phase consisting of acetonitrile and water (containing 5 mmol ammonium acetate adjusted to pH 5.0) within 4 min. The quantification was performed by multiple reaction monitoring with the transitions of m/z 336.0-320.1 for berberine, m/z 352.0-336.1 for palmatine, m/z 338.0-322.1 for jatrorrhizine, m/z 336.0-320.1 for epiberberine, m/z 320.0-292.1 for coptisine and m/z 356.4-192.1 for IS. ResultsThe lower limit of quantification for five protoberberine-type alkaloids was 0.05 ng/mL. Linearity, accuracy, precision, stability and matrix effect of five analytes were all satisfactory. Five protoberberine-type alkaloids were quickly distributed in the brain. Moreover, significant differences in the principal pharmacokinetic parameters such as AUC and T1/2 of the analytes were observed between two groups. ConclusionThe LC-MS/MS method combinated with microdialysis is useful in the brain pharmacokinetic study of five protoberberine-type alkaloids. The results indicated that the rates of analytes absorption in insomnic rats were significantly higher than those in normal rats. Besides, the protoberberine-type alkaloids could bring a direct effect on the neuron in the hippocampus.

Evaluation of the targeted delivery of 5-fluorouracil and ascorbic acid into the brain with ultrasound-responsive nanobubbles

Recently, ultrasound-induced drug delivery into the brain using bubble formulations has been developed. After the brain delivery, however, the information on pharmacokinetics of hydrophilic drugs in the brain is lacking. In this study, to clarify the time-course pharmacokinetics of hydrophilic drugs, we used a brain microdialysis method. Using ultrasound-responsive nanobubbles (bubble liposomes (BLs)) with ultrasound irradiation, two hydrophilic drugs, 5-fluorouracil (5-FU) and ascorbic acid, were delivered into the brain of mice and rats and their time-course pharmacokinetics were evaluated with microdialysis. The results indicated that the time-course pharmacodynamics of ascorbic acid evaluated by examining its antioxidant capacity supported the time-course pharmacokinetics. Additionally, to strengthen the evidences of our evaluation, we varied the effect of BLs dose and duration and intensity of ultrasound irradiation on drug delivery. Among them, when the dose of BLs was changed, the trend of 5-FU intracerebral migration was consistent with other report. In conclusion, we succeeded in clarifying the time-course pharmacokinetics of the two hydrophilic drugs after the brain delivery with bubble formulations and ultrasound irradiation using mice and rats.

Effects of dexmedetomidine, midazolam, and propofol on acetylcholine release in the rat cerebral cortex in vivo

Acetylcholine plays an important role as a neurotransmitter in the central nervous system with involvement in both sleep and arousal. Dexmedetomidine, midazolam, and propofol are widely used for sedation of patients in intensive care medicine. In this study, we have examined the effect of continuous administration of dexmedetomidine, midazolam, and propofol on acetylcholine release in the rat cerebral cortex, using an in vivo microdialysis technique. Following infusion of a control solution, male Wistar rats (n = 6/group) were administered dexmedetomidine at 0.3 μg/kg/min, midazolam at 20 mg/kg/h, or propofol at 50 mg/kg/h over a 2-h period. Using a brain microdialysis method, extracellular acetylcholine concentrations were measured up to 2 h after administration of each agent at 15-min intervals. In the midazolam group, acetylcholine levels were significantly reduced with midazolam infusion, remaining low even after the drug was stopped. In the propofol group, acetylcholine levels were significantly decreased during propofol infusion, but returned to control levels once the infusion was stopped. Dexmedetomidine administration decreased acetylcholine release, but this finding was not statistically significant. From this study, midazolam and propofol but not dexmedetomidine significantly suppressed acetylcholine release in the cerebral cortex at sedative doses. Even though the righting reflex recovered almost the same after the cessation of drug administration, midazolam suppressed acetylcholine release longer than propofol.

Effects of intraperitoneally injected lithium, imipramine and diazepam on nitrate levels in rat amygdala

Nitric oxide (NO) has been studied in relation to the etiologies of various neurologic and psychiatric diseases. However, little is known about whether clinically available psychotropic drugs affect the NO system in the brain. Using an in vivo brain microdialysis method, the effects of intraperitoneally administered lithium, imipramine and diazepam on levels of , a marker of in vivo NO production, were investigated in the rat amygdala. Lithium significantly reduced, while imipramine raised, levels as compared with controls. These observations suggest that lithium and imipramine induce opposite effects on NO-related systems in the brain.

L-DOPA-derived extracellular dopamine in the striatum with dopaminergic denervation: role of serotonergic neurons in l-DOPA metabolism

In order to determine the state of dopamine (DA) derived from exogenously administered l-3,4-dihydroxyphenylalanine (l-DOPA) in the striatum of Parkinson's disease, we have applied an in vivo brain microdialysis method to rats with dopaminergic denervation with 6-hydroxydopamine (6-OHDA), the rat model of Parkinson's disease, and measured extracellular DA levels in the striatum after l-DOPA administration. We obtained the following results: (1) Inhibitory effect of the potent D2 agonist quinpirole on l-DOPA-derived extracellular DA was not detected in the denervated striatum; (2) reserpine pretreatment greatly reduced l-DOPA-derived extracellular DA; (3) additional serotonergic denervation to rats with dopaminergic denervation induced a significant reduction in l-DOPA-derived extracellular DA; and (4) activation of 5-hydroxytryptamine (serotonin) (5-HT)1A receptors attenuated an increase in l-DOPA-derived extracellular DA. These results suggest that the conversion of exogenous l-DOPA to DA and the storage and release of converted DA occur in serotonergic nerve terminals when dopaminergic neurons are denervated. It is possible that serotonergic neurons play a critical role in the therapeutic effect of l-DOPA in Parkinson's disease.

Effects of phencyclidine on behavior and extracellular levels of dopamine and its metabolites in neonatal ventral hippocampal damaged rats.

The use of neonatal hippocampal lesioned rats is well established in animal models of schizophrenia. Moreover, the dysfunction of N-methyl-D-aspartate (NMDA) neurotransmission may play a crucial role in the pathophysiology of schizophrenia. To examine the role of NMDA neurotransmission in the neonatal ventral hippocampal damaged rat. In initial experiments, we compared the effects of mild environmental stress (HAB) and the injection of saline and methamphetamine (MAP, 1.5 mg/kg, IP) in lesioned and sham-operated rats. We also examined the effects of a single injection of phencyclidine (PCP, 10 mg/kg, IP) on locomotor activity and extracellular levels of dopamine (DA) and its metabolites in the nucleus accumbens (NAc) of lesioned and sham-operated rats using an in vivo brain microdialysis method. Compared with sham-operated controls, the lesioned rats showed increased locomotor activity at postnatal day 56 (PD56) but not at PD35 after HAB and MAP administration. Similarly, the lesioned rats showed increased locomotor activity at PD56 but not at PD35 after PCP administration. Unexpectedly, the increase in DA levels was significantly greater in the sham-operated rats than in the lesioned rats. The results suggest that neonatal hippocampal lesioned rats are accompanied by the dysfunction of NMDA neurotransmission. They also suggest that hyperresponsiveness to PCP following neonatal hippocampal lesions does not depend on the extracellular DA concentration in the NAc.

実験的こう合障害がラット前頭皮質ドーパミン放出に及ぼす影響

It is very interesting to determine whether disturbance of mastication following occlusal dysfunction has an effect on mental stress. Therefore, an experimental occlusal interference was attached, and the effect on dopamine release from the rat prefrontal cortex was examined.The extracellular fluid dopamine volume of the prefrontal cortex was measured by the brain microdialysis method.1. Though experimental occlusal interference during nonfunctional periods was intraoral, it. was found that the metabolism of the prefrontal cortex dopamine did not change.2. Experimental occlusal interference during functional periods, that is, feeding periods with cubed food under occlusal interference increased the prefrontal cortex dopamine volume. It was clarified that experimental occlusal interference strongly impacts the affect of the rat by changing the metabolism of dopamine in the prefrontal cortex.3. Cubed food and powdered food increased the prefrontal cortex dopamine volume in the same way. It was found that there was no difference in transition of the increase in the prefrontal cortex dopamine volume from the difference in the texture of the food.Experimental occlusal interference related to feeding behavior affected dopamine metabolism in the rat prefrontal cortex, and it was clarified that a strong stress reaction was induced by experimental occlusal interference.

Synthesis of 4(5)-[5-(Aminomethyl)tetrahydrofuran-2-yl- or 5-(Aminomethyl)-2,5-dihydrofuran-2-yl]imidazoles by Efficient Use of a PhSe Group: Application to Novel Histamine H3-Ligands1

(+)-4(5)-[(2R,5S)-(5-Aminomethyl)tetrahydrofuran-2-yl]imidazole 1 and its C2‘ epimer (−)-2, which are the 5‘-amino derivatives of a novel imidazole C-nucleoside, were synthesized via β- and α-2‘-phenylselenenyl nucleosides 15 and 16. The anomers 15 and 16 were provided by a new synthetic method for C-nucleosides via the elimination of PhSeCl and selenocyclization from diol intermediates 12 and 14, starting from l-glutamic acid. Their ent-1 and ent-2 (imifuramine), the latter of which was indicated as a novel type of histamine H3-agonist confirmed by an in vivo brain microdialysis method, were synthesized by the same methodology from d-glutamic acid. The four isomers (3, 4, ent-3, and ent-4) of a 4(5)-[(5-aminomethyl)-2,5-dihydrofuran-2-yl]imidazole were also synthesized via the oxidative elimination of the PhSe group of the key intermediates (15, 16, ent-15, and ent-16). In connection with this study, 4(5)-(5-aminomethylfuran-2-yl)-1H-imidazole (5) was also synthesized starting from d-ribose.

Ischemia-induced glutamate release in the dentate gyrus. A microdialysis study in the gerbil

The levels of extracellular glutamate were measured in the dentate gyrus by using an in vivo brain microdialysis method to determine whether the ischemia-induced glutamate release might be correlated with the neuronal vulnerability to ischemia. A microdialysis membrane was placed in CA4 (vulnerable to ischemia) and the molecular and granule cell layers of the dentate gyrus (resistant to ischemia) of gerbils. A significant increase in glutamate levels was induced in the normal dentate gyrus during 10-min ischemia. The increase was completely suppressed during the first 5 min of ischemia when CA4 neurons were eliminated. Thus, it was indicated that during the first 5 min of ischemia glutamate was released mostly from CA4 neurons but not from granule cells of the dentate gyrus. During the second half of 10-min ischemia, a significant increase in glutamate release was induced even in the dentate gyrus where CA4 neurons were eliminated; this increase was significantly suppressed by inhibiting proliferation of astrocytes. A large part of glutamate that was released during the second half of 10-min ischemia was considered to be attributable to glutamate release from astrocytes.

Effect of propranolol on central neurotransmitter release in Wistar rats analysed by brain microdialysis.

1. The effect of propranolol on amino acid neurotransmitter release in the rostral ventrolateral medulla (RVLM) was examined in urethane-anaesthetized male Wistar rats. 2. Amino acids released in the RVLM in response to intravenous administration of propranolol (0.4 mg/kg per min; n = 6) or nitroglycerin (0.02 mg/kg per min; n = 5) were determined by the brain microdialysis method. 3. Amino acids in dialysates were analysed by high-performance liquid chromatography and were quantified by ultraviolet absorbance. 4. Administration of both intravenous propranolol and nitroglycerin significantly decreased arterial blood pressure. Heart rate was decreased only by propranolol. 5. The reduction in arterial blood pressure produced by intravenous propranolol was accompanied by a decrease in the release of the excitatory amino acid glutamate in the RVLM. 6. The reduction in arterial blood pressure following intravenous nitroglycerin was not accompanied by a release of glutamate. 7. There were no significant changes in the levels of other amino acids (glycine, taurine, GABA) following either propranolol or nitroglycerin. 8. The decrease in glutamate release in the RVLM may account, in part, for the central depressor mechanism of propranolol.

Synthesis of imifuramine and its stereoisomers exhibiting histamine H 3-agonistic activity

The four possible stereoisomers of a novel imidazole C-nucleoside derivative were synthesized by the efficient use of a PhSe group. Among them, (+)-4(5)-[(2R,5R)-5-(aminomethyl)tetrahydrofuran-2-yl]imidazole (imifuramine) was indicated as a novel type of histamine H 3-agonist by a brain microdialysis method.

Nociceptive stimulus induces release of endogenous β-endorphin in the rat brain

The hypothesis that the naturally occurring analgesic peptide, β-endorphin, is released in the brain in response to pain had never been directly validated. In this study, we applied a brain microdialysis method for monitoring β-endorphin release in vivo, to test this hypothesis in the brains of conscious, freely moving rats. Herein we first show that endogenous β-endorphin can be measured in vivo in the brain under physiological conditions. Upon induction of a nociceptive stimulus by injection of formalin into the hind-paws of rats, the extracellular levels of β-endorphin in their arcuate nucleus increased by 88%, corresponding to their nociceptive response. This direct evidence for the release of endogenous β-endorphin in the brain in response to nociceptive stimulus indicates a possible mechanism for organisms to cope with pain.

ESR imaging of the rat brain with a nitroxide radical perfused by in vivo microdialysis

We report here our investigation of the spatial distribution of free radicals using an electron spin resonance (ESR)-imaging system combined with an in vivo brain microdialysis method, which was performed in the resonator of the ESR-imaging system. A nonmagnetic cannula, newly developed in this study, was used for the perfusion of the exogenous free radicals agent. A nitroxide, 3-carbamoyl-2,2,5,5-tetramethylpyrrolidine-1-oxyl (carbamoyl PROYXL), was used as the imaging agent in saline solution at a concentration of 0.3 M, which was perfused into the right caudate putamen of the rat at 2 μl/min by a microinfusion pump. Two-dimensional ESR projection of the Z- X plane, which was clearly distinguished (about φ 10 mm) from the nonperfused brain area, was obtained 6 h after the beginning of perfusion of carbamoyl PROXYL. The present method is considered to be a useful tool to introduce stable free radicals into a specific area of the brain.

Ipsapirone enhances the dopamine outflow via 5-HT 1A receptors in the rat prefrontal cortex

In the present study, we investigated both the effect of ipsapirone on the dopamine outflow and its selectivity towards 5-HT 1A receptors in the rat prefrontal cortex. Using a brain microdialysis method in freely moving animals, it was found that ipsapirone, 5 and 10 mg/kg dose-dependently enhanced the outflow of dopamine, while 2.5 mg/kg was ineffective. The above effects of ipsapirone were mimicked by buspirone (2.5 and 5 mg/kg), another 5-HT 1A receptor agonist, but not 1-PP (1-pyrimidinylpiperazine, 5 mg/kg) — a centrally active metabolite of ipsapirone. The effect of ipsapirone (10 mg/kg) on the dopamine outflow in the rat prefrontal cortex was antagonized by 1-(2-methoxyphenyl)-4-[4-(2-phthalimido)butyl]piperazine (NAN-190, 1 mg/kg) and ( N-tert-butyl-3-(4-(2-methoxyphenylpiperazin-1-yl)-2-phenylpropionamide (WAY 100135, 10 mg/k.g.), i.e. substances with agonistic/antagonistic and antagonistic properties in relation to 5-HT 1A receptors, respectively. NAN-190 (1 mg/kg) enhanced the outflow of dopamine, while WAY 100135 (10 mg/kg) failed to alter it. It is concluded that 5-HT 1A receptor agonists may be involved in the regulation of dopaminergic neurotransmission in the rat prefrontal cortex and may have therapeutic potential in the treatment of disorders associated with dysfunction of the mesocortical dopaminergic system.

Role of dopaminergic neuronal system in dizocilpine-induced acetylcholine release in the rat brain

The effects of dopaminergic receptor antagonists on dizocilpine-induced increase in extracellular acetylcholine (ACh) levels in the rat parietal cortex were examined in freely-moving rats, using an in vivo brain microdialysis method. Dizocilpine (0.5 mg/kg) significantly increased extracellular ACh levels in the rat parietal cortex and hippocampus, but not in the striatum. Pretreatment with alpha-methyl-p-tyrosine methyl ester (alpha MpT) delayed the onset but prolonged the duration of the dizocilpine-induced increases in extracellular ACh levels. The dopamine D2 receptor antagonist, haloperidol, showed dual effects similarly to alpha MpT, while the dopamine D1 receptor antagonist, SCH23390, prolonged, but did not delay, the onset of the dizocilpine-induced increases in ACh levels. These results suggest that the dopaminergic system is involved in the dizocilpine-induced increase in the extracellular ACh level in the parietal cortex in two ways, through both dopamine D1 and D2 receptors.

Dysfunction of cholinergic and dopaminergic neuronal systems in beta-amyloid protein--infused rats.

Accumulations of beta-amyloid protein are characteristic and diagnostic features of the brain of Alzheimer's disease patients; however, the physiological role of this protein in CNS is unknown. We have previously reported that continuous infusion of beta-amyloid protein into rat cerebral ventricle impairs learning ability and decreases choline acetyltransferase activity, a marker enzyme of cholinergic neuron. In this study, the effects of beta-amyloid protein infusion on the release of neurotransmitters in cholinergic and dopaminergic neuronal systems were investigated by using an in vivo brain microdialysis method. Nicotine-stimulated release of acetylcholine and dopamine in these animals was significantly lower than that in vehicle-infused rats. Further, dopamine release induced by high-K stimulation was decreased in beta-amyloid protein-infused rats compared with vehicle-infused rats. These results suggest that the release of the two transmitters, acetylcholine and dopamine, was decreased by beta-amyloid protein and that learning deficits observed in the beta-amyloid protein-infused rats are partly due to the impairment of neurotransmitter release. Furthermore, continuous infusion of beta-amyloid protein may be a useful method to produce the animal model of Alzheimer's disease.