Transfection of an active cytochrome P450 arachidonic acid epoxygenase indicates that 14,15-epoxyeicosatrienoic acid functions as an intracellular second messenger in response to epidermal growth factor

A common feature of most isolated cell systems is low or undetectable levels of bioactive cytochrome P450. We therefore developed stable transfectants of the renal epithelial cell line, LLCPKcl4, that expressed an active regio- and enantioselective arachidonic acid (AA) epoxygenase. Site-specific mutagenesis was used to convert bacterial P450 BM-3 into an active regio- and stereoselective 14S,15R-epoxygenase (F87V BM-3). In clones expressing F87V BM-3 (F87V BM-3 cells), exogenous AA induced significant 14S,15R-epoxyeicosatrienoic acid (EET) production (241. 82 ng/10(8) cells, >97% of total EETs), whereas no detectable EETs were seen in cells transfected with vector alone. In F87V BM-3 cells, AA stimulated [3H]thymidine incorporation and increased cell proliferation, which was blocked by the tyrosine kinase inhibitor, genistein, by the phosphatidylinositol 3 (PI-3) kinase inhibitors, wortmannin and LY294002, and by the mitogen-activated protein kinase kinase inhibitor, PD98059. AA also induced tyrosine phosphorylation of extracellular signal-regulated kinase (ERK) and PI-3 kinase that was inhibited by the cytochrome P450 BM-3 inhibitor, 17-ODYA. Epidermal growth factor (EGF) increased EET production in F87V BM-3 cells, which was completely abolished by pretreatment with either 17-ODYA or the phospholipase A2 (PLA2) inhibitor, quinacrine. Compared with vector-transfected cells, F87 BM-3 transfected cells demonstrated marked increases in both the extent and sensitivity of DNA synthesis in response to EGF. These changes occurred in the absence of significant differences in EGF receptor expression. As seen with exogenous AA, EGF increased ERK tyrosine phosphorylation to a significantly greater extent in F87V BM-3 cells than in vector-transfected cells. Furthermore, in these control cells, neither 17-ODYA nor quinacrine inhibited EGF-induced ERK tyrosine phosphorylation. On the other hand, in F87V BM-3 cells, both inhibitors reduced ERK tyrosine phosphorylation to levels indistinguishable from that seen in cells transfected with vector alone. These studies provide the first unequivocal evidence for a role for the AA epoxygenase pathway and endogenous EET synthesis in EGF-mediated signaling and mitogenesis and provide compelling evidence for the PLA2-AA-EET pathway as an important intracellular-signaling pathway in cells expressing high levels of cytochrome P450 epoxygenase.     

Blockade of cerebral blood flow response to insulin-induced hypoglycemia by caffeine and glibenclamide in conscious rats

The possibility that adenosine and ATP-sensitive potassium channels (KATP) might be involved in the mechanisms of the increases in cerebral blood flow (CBF) that occur in insulin-induced hypoglycemia was examined. Cerebral blood flow was measured by the [14C]iodoantipyrine method in conscious rats during insulin-induced, moderate hypoglycemia (2 to 3 mmol/L glucose in arterial plasma) after intravenous injections of 10 to 20 mg/kg of caffeine, an adenosine receptor antagonist, or intracisternal infusion of 1 to 2 mumol/L glibenclamide, a KATP channel inhibitor. Cerebral blood flow was also measured in corresponding normoglycemic and drug-free control groups. Cerebral blood flow was 51% higher in untreated hypoglycemic than in untreated normoglycemic rats (P < 0.01). Caffeine had a small, statistically insignificant effect on CBF in normoglycemic rats, but reduced the CBF response to hypoglycemia in a dose-dependent manner, i.e., 27% increase with 10 mg/kg and complete elimination with 20 mg/kg. Chemical determinations by HPLC in extracts of freeze-blown brains showed significant increases in the levels of adenosine and its degradation products, inosine and hypoxanthine, during hypoglycemia (P < 0.05). Intracisternal glibenclamide had little effect on CBF in normoglycemia, but, like caffeine, produced dose-dependent reductions in the magnitude of the increases in CBF during hypoglycemia, i.e., +66% with glibenclamide-free artificial CSF administration, +25% with 1 mumol/L glibenclamide, and almost complete blockade (+5%) with 2 mumol/L glibenclamide. These results suggest that adenosine and KATP channels may play a role in the increases in CBF during hypoglycemia.     

Stereoselective epoxidation of arachidonic acid by cytochrome P-450s 2CAA and 2C2

In the present study, we determined the stereoselective epoxidation of arachidonic acid by cytochrome P-450 (P-450) 2CAA and P-450 2C2, two arachidonic acid epoxygenases found in rabbit renal cortex, by chiral normal-phase high-performance liquid chromatography (HPLC)-analysis. Purified P-450 2CAA reconstituted with P-450 oxidoreductase, lipid and cytochrome b5 or microsomes isolated from COS-1 cells expressing P-450 2C2 were incubated in the presence of [1-14C]arachidonic acid. The epoxide metabolites 14,15- and 11,12-epoxyeicosatrienoic acids (EETs) were purified by reverse-phase HPLC and derivatized to methyl (14,15-EET) and pentafluorobenzyl (11,12-EET) esters. Enantiomers of 14,15-EET-methyl ester and 11,12-EET-pentafluorobenzyl ester were resolved on Chiralcel OB and OD columns, respectively, with a mobile phase of 0.15% 2-propanol in n-hexane. P-450 2CAA and P-450 2C2 produce 11,12- and 14,15-EETs in distinct ratios but are equally stereoselective at the 11,12-position. P-450 2CAA produced 11(S), 12(R)-EET with 63% stereoselectivity, and P-450 2C2 produced the same enantiomer with 61% stereoselectivity. Both enzymes are also stereoselective at the 14,15- position, preferentially producing the 14(R), 15(S)-EET. P-450 2CAA produces this enantiomer with 72% selectivity, and P-450 2C2 produces it with 62% selectivity. The results of this study indicate that P-450 2CAA and P-450 2C2 are not only regioselective but also exhibit a high degree of stereoselectivity.     

Role of metabolism by esterase and cytochrome P-450 in cocaine-induced suppression of the antibody response

To investigate the role of metabolism in cocaine-induced immunosuppression, diazinon and beta-ionone were administered as an esterase inhibitor and a cytochrome P-450 (P-450) inducer, respectively, to B6C3F1 female mice. When 10 or 30 mg/kg of diazinon was administered 30 min before cocaine (30 mg/kg) was administered i.p. for 7 consecutive days, the suppression of the T-dependent antibody response to sheep red blood cells was potentiated greatly when compared to the suppression by cocaine alone. Spleen and thymus weights were decreased significantly and serum glutamate-pyruvate transaminase activities were elevated dramatically when cocaine and diazinon were administered together. beta-Ionone was administered s.c. for 7 consecutive days and the P-450 activities were determined 3 days after the last administration. beta-Ionone induced cocaine N-demethylation, which is the first step in the activation of cocaine to the metabolites capable of producing hepatotoxicity, as well as P-450IA1- and P-450IIB1-specific monooxygenases. The inductive effects of beta-ionone on P-450IA1/2 and P-450IIB1/2 proteins were confirmed by using Western immunoblotting with selective monoclonal antibodies. In addition, when beta-ionone (600 mg/kg) was administered with cocaine for 7 days, the suppression of the antibody response was potentiated greatly, thymus weight was decreased significantly and serum glutamate-pyruvate transaminase was elevated. Our present results suggest that inhibition of the esterase pathway of cocaine shunts the metabolism of cocaine into an immunotoxic pathway, and that the metabolism of cocaine by P-450 may be the critical pathway for the generation of the metabolites capable of suppressing the antibody response.     

Hypercapnic cerebral blood flow in spontaneously hypertensive rats

OBJECTIVE: Hypercapnic cerebral vasodilation appears to be endothelium-dependent, as it involves nitric oxide and prostaglandins. Since chronic hypertension has been associated with impaired endothelial function, we designed a study to find out whether hypercapnic cerebral blood flow and its nitric oxide- and prostaglandin-sensitive component is reduced in spontaneously hypertensive rats (SHR) compared with normotensive controls. METHODS: Cerebral blood flow was measured in enflurane-anesthetized SHR (n=53), Wistar-Kyoto (WKY, n=20) and Sprague-Dawley (n=50) rats using the hydrogen clearance method. Cerebral blood flow was measured during eucapnia and hypercapnia; it was also assessed after administering either nonisoform-selective or isoform-selective neuronal nitric oxide synthase inhibitors and during inhibition of prostaglandin production. RESULTS: Hypercapnic cerebral blood flow did not differ among the strains. Nitric oxide synthase inhibition with intracortical N(G)-monomethyl-L-arginine reduced hypercapnic cerebral blood flow in SHR by 23+/-4% and in Sprague-Dawley rats by 23+/-7% without affecting eucapnic flow. Intraperitoneal administration of the inhibitor of neuronal nitric oxide synthase, 7-nitroindazole, reduced eucapnic flow by 18+/-5% in SHR and 27+/-5% in WKY rats, and hypercapnic flow by 48+/-3 and by 51+/-6%, respectively. Indomethacin produced a similar decrease in hypercapnic flow in Sprague-Dawley rats and SHR (49+/-5 and 62+/-4%, respectively). CONCLUSION: Hypercapnic cerebral blood flow was not impaired in SHR. The contribution of nitric oxide- and prostaglandin-dependent vasodilation appeared to be intact Our results are consistent with the hypothesis that neuronal rather than endothelial production of nitric oxide may be responsible for maintaining hypercapnic cerebral vasodilation in SHR.     

Diminished regional cerebral blood flow response to vibration in patients with blepharospasm

Mutations of the human Patched gene ( PTCH ) have been identified in individuals with the nevoid basal cell carcinoma syndrome (NBCCS) as well as in sporadic basal cell carcinomas and medulloblastomas. We have isolated a homologue of this tumour suppressor gene and localized it to the short arm of chromosome 1 (1p32.1-32.3). Patched 2 ( PTCH2 ) comprises 22 coding exons and spans approximately 15 kb of genomic DNA. The gene encodes a 1203 amino acid putative transmembrane protein which is highly homologous to the PTCH product. We have characterized the genomic structure of PTCH2 and have used single-stranded conformational polymorphism analysis to search for mutations in PTCH2 in NBCCS patients, basal cell carcinomas and in medulloblastomas. To date, we have identified one truncating mutation in a medulloblastoma and a change in a splice donor site in a basal cell carcinoma, suggesting that the gene plays a role in the development of some tumours.     

Role of nitric oxide in angiotensin IV-induced increases in cerebral blood flow

Whether a rapid elevation of serum gliclazide concentration in human subjects can be achieved through an acceleration of dissolution of gliclazide from a formulation was examined. A soft gelatin capsule containing PEG 400, PEG 4000, Tween 20 and glycerin was prepared as a formulation that may accelerate dissolution of gliclazide. The in vitro dissolution of gliclazide at pH 7.2 was identical for the soft capsule and conventional tablets, Diamicron and Diberin. However, at pH 1, 2 and 4.0 the dissolution from the soft capsule was more rapid compared to the tablets. When bioavailability parameters were compared following oral administration of the soft capsule and Diamicron to 16 healthy Korean male subjects, the parameters representing the amount of adsorption (i.e. the area under the serum gliclazide concentration vs. time curve up to 24 h, AUC24, and the peak serum concentration Cmax) were not statistically different for both formulations. However, the time required to reach the peak (Tmax) was significantly shorter for the soft capsule than for the Diamicron. Our results, therefore, indicate that a rapid elevation of serum gliclazide concentration following oral administration of a formulation can be achieved by accelerating the in vitro dissolution of gliclazide from the formulation into the acidic buffers. Thus, the rate of gastrointestinal absorption of gliclazide appears to be dependent on its in vivo dissolution rate in gastric fluid. A soft capsule containing a PEG 400 suspension of gliclazide appears to be an appropriate formulation for accelerating the dissolution.     

Reduction of regional cerebral blood flow by melatonin in young rats

Melatonin (10 ng) was subcutaneously administered to 14-day-old Sprague-Dawley rats. Regional blood flow (rCBF) was measured in 22 anatomically defined structures 20 min later using iodo[14C]antipyrine and quantitative autoradiography. rCBF was markedly reduced in the cerebral areas supplied by circle of Willis and the basilar arteries. Melatonin also significantly decreased blood flow to choroid plexuses. These findings suggest that circulating melatonin may contribute to regulation of cerebral blood flow and brain fluid balance.     

The effects of sevoflurane on cerebral blood flow autoregulation in rats

In all species tested, except humans, biological differences between vitamins D2 and D3 are accepted as fact. To test the presumption of equivalence in humans, we compared the ability of equal molar quantities of vitamin D2 or D3 to increase serum 25-hydroxyvitamin D [25(OH)D], the measure of vitamin D nutrition. Subjects took 260 nmol (approximately 4000 IU) vitamin D2 (n=17) or vitamin D3 (n=55) daily for 14 d. 25(OH)D was assayed with a method that detects both the vitamin D2 and D3 forms. With vitamin D3, mean (+/-SD) serum 25(OH)D increased from 41.3+/-17.7 nmol/L before to 64.6+/-17.2 nmol/L after treatment. With vitamin D2, the 25(OH)D concentration went from 43.7+/-17.7 nmol/L before to 57.4+/-13.0 nmol/L after. The increase in 25(OH)D with vitamin D3 was 23.3+/-15.7 nmol/L, or 1.7 times the increase obtained with vitamin D2 (13.7+/-11.4 nmol/L; P=0.03). There was an inverse relation between the increase in 25(OH)D and the initial 25(OH)D concentration. The lowest 2 tertiles for basal 25(OH)D showed larger increases in 25(OH)D: 30.6 and 25.5 nmol/L, respectively, for the first and second tertiles. In the highest tertile [25(OH)D >49 nmol/L] the mean increase in 25(OH)D was 13.3 nmol/L (P < 0.03 for comparison with each lower tertile). Although the 1.7-times greater efficacy for vitamin D3 shown here may seem small, it is more than what others have shown for 25(OH)D increases when comparing 2-fold differences in vitamin D3 dose. The assumption that vitamins D2 and D3 have equal nutritional value is probably wrong and should be reconsidered.     

Relationship between cytochrome P-450 induction by rifampicin, hepatic volume and portal blood flow in man

OBJECTIVE: Induction of hepatic cytochrome P-450-dependent oxidative metabolism is related to an almost identical increase (30%) in both the liver weight and portal blood flow in animals. In humans by contrast, an increased liver blood flow (44%) but no significant increase in liver volume has been reported. DESIGN: Therefore, we studied prospectively the relationship between P-450 induction by rifampicin, hepatic volume and portal blood flow in 10 healthy volunteers. METHODS: After a pre-treatment phase (day 1 to 7) the 10 volunteers received 600 mg/day of rifampicin from day 7 to 12. The urinary 6-beta-hydroxycortisol output as a measure of oxidative metabolism (CYP3A4) and portal blood flow (pulsed Doppler ultrasound) were determined on days 1, 7, 11 and 13. Hepatic magnetic resonance volumetry was performed on days 1 and 13. RESULTS: Urinary 6-beta-hydroxycortisol output increased in all volunteers (P = 0.0051) from a median of 2.15 micrograms/day/kg (1.8-3.3 micrograms/day/kg) on day 1 to 9.9 micrograms/day/kg (5.7-14 micrograms/day/kg) on day 13. In 9 of 10 volunteers induction by rifampicin was related to an increase (P = 0.0218) in liver volume from a median of 1570 cm3 (1390-1830 cm3) to a median of 1690 cm3 (1420-1860 cm3). The portal flow as assessed by colour Doppler ultrasound did not change significantly between day 1 (median 22 cm/s (15-35 cm/s)) and day 13 (median 19 cm/s (16-39 cm/s)). CONCLUSION: A fourfold increase of urinary 6-beta-hydroxycortisol output after induction of cytochrome P-450 by rifampicin is associated with a significant but less than 10% increase in human liver volume. No increase of portal perfusion as assessed by Doppler ultrasound could be detected in this study.     

Ocular and regional cerebral blood flow in aging Fischer-344 rats

Vascular remodeling and changes in vascular responsiveness occur in the rat cerebrum with old age. This includes reductions in cerebral arteriolar numerical density, cross-sectional area, distensibility, the relative proportion of distensible elements in the cerebral arteriolar wall, and reduced endothelium-dependent relaxation. The purpose of this study was to test the hypothesis that old age results in an increase in vascular resistance and, correspondingly, a decrease in blood flow to ocular, regional cerebral, and spinal tissue in the rat. Blood flow was measured in the eye, olfactory bulb, left and right cerebrum, pituitary gland, midbrain, pons, cerebellum, medulla, and spinal cord of juvenile (2-mo-old, n = 6), adult (6-mo-old, n = 7), and aged (24-mo-old, n = 7) male Fischer-344 rats. Arterial pressure and blood flow were used to calculate vascular resistance. Vascular resistance in the eye of aged rats (6.03 +/- 1.08 mmHg . ml-1 . min . 100 g) was higher than that in juvenile (3.83 +/- 0.38 mmHg . ml-1 . min . 100 g) and adult rats (3.12 +/- 0.24 mmHg . ml-1 . min . 100 g). Similarly, resistance in the pons of older rats (2.24 +/- 0.55 mmHg . ml-1 . min . 100 g) was greater than in juvenile (0.66 +/- 0.06 mmHg .ml-1 . min . 100 g) and adult rats (0.80 +/- 0.11 mmHg . ml-1 . min . 100 g). In contrast, vascular resistance in the pituitary gland was lower in the aged rats (juvenile, 3.09 +/- 0.22; adult, 2.79 +/- 0.42; aged, 1.73 +/- 0.32 mmHg . ml-1 . min . 100 g, respectively). Vascular resistance was not different in other cerebral tissues or in the spinal cord in the aged rats. These data suggest that regional cerebral and spinal blood flow and vascular resistance remain largely unchanged in conscious aged rats at rest but that elevations in ocular vascular resistance and, correspondingly, decreases in ocular perfusion with advanced age could have serious adverse effects on visual function.     

Long-term prognosis in stroke related to cerebral blood flow

In 180 patients, cerebral blood flow was measured between one and six weeks after acute cerebrovascular ischemia. Patients were grouped according to their clinical deficits at the time of blood flow study. Statistical analysis revealed highly significant differences among the flow values of the groups. The patients were followed up for a period of up to 6 years (mean, 34 months) after the attack, and then regrouped. In these groups, the statistical significances of differences among the flow values was even higher. Patients also were grouped according to changes in neurologic status, and again the flow values differed significantly among the groups. The results show significant relationships among flow values after cerebrovascular ischemia and neurologic deficits, change in status, and the final functional state. Cerebral blood flow measurement together with other clinical signs permits an estimation of a patient's chance for functional recovery after a stroke.     

Forearm blood flow in response to stress.

Examined the relationship between stress, forearm blood flow, and subjective anxiety. In 26 normal male Ss, electric shocks applied to the opposite forearm led to a rapid rise in anxiety and forearm blood flow. With repeated regular shocks, habituation of the vasomotor response occurred and anxiety became less intense. Changes appeared to be related to S's ability to predict accurately the intensity and frequency of the stimulus. Mental arithmetic under harassment produced a more gradual but greater rise in blood flow without an equal rise in anxiety. 23 alcoholic Ss who were judged to have limited motivation and ego strength showed a significantly lower vasomotor response during mental arithmetic. It is concluded that changes in forearm blood flow observed during stress cannot be related only to an increase in anxiety. This response seems to depend also on the significance of the stimulus to S, his psychiatric status, and his level of motivation. (19 ref.) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Prognostic behavioral criteria and the characteristics of the cerebral blood flow in rats with different sensitivities to emotional stress

In anesthetised Wistar adult male rats, a model of circulatory brain ischemia was created. A behavioural difference as well as a difference in normal focal cerebral blood flow (CBF), were revealed between rats resistant against and predisposed to emotional stress. The mortality level was 10% in former rats and 27% in latter ones.     

Role of substance P in pressor response of central amygdaloid nucleus to glutamate

Substance P (SP)-immunoreactive cells and the axon terminals are widely distributed in the central amygdaloid nucleus (AC) and its important projection areas. The present study showed that (1) Excitation of the AC by glutamate (Glu) or injection of SP into the AC projection areas: locus coeruleus (LC), nucleus parabrachialis (NPB), periaqueductal gray matter (PAG) or lateral hypothalamus-perifornical region (LH/PF), all elicited pressor response. (2) Preinjection of DPDPDT (a SP antagonist) into bilateral LC, NPB, PAG or LH/PF could attenuate the AC pressor response to Glu. (3) Intra-RVLM (rostral ventrolateral medulla) preinjection of either phentolamine, propranolol or atropine (but not GDEE, a Glu antagonist) could also reduce the AC pressor response. Taken together with our previous findings that the alpha-, beta-, M-receptors in RVLM mediated the pressor response to LC excitation, alpha-receptors mediated the NPB pressor response, alpha- and beta-receptors mediated the PAG pressor response; these results indicate that the SPergic projections of the AC not only directly act upon the brainstem pressor areas (LC, NPB, PAG)-RVLM system, but also indirectly via the LH/PF act upon the brainstem pressor areas-RVLM system to induce the pressor response.     

Fast and slow components of cerebral blood flow response to step decreases in end-tidal PCO2 in humans

This study examined the dynamics of the middle cerebral artery (MCA) blood flow response to hypocapnia in humans (n = 6) by using transcranial Doppler ultrasound. In a control protocol, end-tidal PCO2 (PETCO2) was held near eucapnia (1.5 Torr above resting) for 40 min. In a hypocapnic protocol, PETCO2 was held near eucapnia for 10 min, then at 15 Torr below eucapnia for 20 min, and then near eucapnia for 10 min. During both protocols, subjects hyperventilated throughout and PETCO2 and end-tidal PO2 were controlled by using the dynamic end-tidal forcing technique. Beat-by-beat values were calculated for the intensity-weighted mean velocity (VIWM), signal power (P), and their instantaneous product (P.VIWM). A simple model consisting of a delay, gain terms, time constants (tauf,on, tauf, off) and baseline levels of flow for the on- and off-transients, and a gain term (gs) and time constant (taus) for a second slower component was fitted to the hypocapnic protocol. The cerebral blood flow response to hypocapnia was characterized by a significant (P < 0.001) slow progressive adaptation in P.VIWM, with gs = 1.26 %/Torr and taus = 427 s, that persisted throughout the hypocapnic period. Finally, the responses at the onset and relief of hypocapnia were asymmetric (P < 0.001), with tauf,on (6.8 s) faster than tauf,off (14.3 s).     

Role of cytochrome P-450 as a source of catalytic iron in cisplatin-induced nephrotoxicity

BACKGROUND: Iron plays a role in free radical-mediated tissue injury, including cisplatin-induced nephrotoxicity. However, the source of iron (catalyzing free radical reactions) is not known. We examined the role of cytochrome P-450 as a source of catalytic iron in cisplatin-induced nephrotoxicity both in vivo and in vitro. METHODS: Cisplatin-induced acute renal failure was produced in rats by intraperitoneal injection of cisplatin (10 mg/kg body wt). Piperonyl butoxide, a cytochrome P-450 inhibitor, was administered intraperitoneally (400 mg/kg body wt twice at 48-hr intervals) prior to cisplatin injection. The effects of cisplatin in the absence or presence of piperonyl butoxide on the belomycin-detectable iron, cytochrome P-450 content in the kidney, and renal functional and histological changes were evaluated. In an in vitro study, the effect of cytochrome P-450 inhibitors, cimetidine or piperonyl butoxide, on cisplatin-induced cytotoxicity and catalytic iron release from LLC-PK1 cells was examined. RESULTS: In cisplatin-treated rats, there was a marked decrease in the cytochrome P-450 content specifically in the kidney, accompanied by increased bleomycin-detectable iron content in the kidney. Piperonyl butoxide prevented cisplatin-induced loss of cytochrome P-450 as well as the increase of bleomycin-detectable iron in the kidney, along with both functional and histological protection. Both cimetidine and piperonyl butoxide prevented cisplatin-induced increase in bleomycin-detectable iron and cytotoxicity in LLC-PK1 cells. Treatment of cimetidine did not affect cellular uptake of cisplatin. CONCLUSION: Cytochrome P-450, a group of heme proteins, may serve as a significant source of catalytic iron in cisplatin-induced nephrotoxicity.     

Regional cerebral blood flow and cerebrospinal fluid amino acid analysis in elderly dementia

Regional cerebral blood flow and amino acid concentration in the cerebrospinal fluid were studied in 12 cases of vascular dementia, 12 cases of Alzheimer's disease, 12 cases of chronic alcoholism, and 12 age-matched healthy controls. In vascular dementia, blood flows were decreased in the cerebral cortex, frontal white matter, thalamus, caudate nucleus, and putamen and alpha-aminobutyric acid and glutamic acid concentrations were increased in the cerebrospinal fluid. In Alzheimer's disease, blood flows were decreased in the frontal cortex, parietal cortex, temporal cortex, and frontal white matter and alanine concentration was increased in the cerebrospinal fluid. In chronic alcoholism, blood flows were decreased in the cerebral cortex, thalamus, and putamen and urea, alanine, and glycine concentrations were increased in the cerebrospinal fluid.     

Role of nitric oxide in maintenance of basal anterior choroidal blood flow in rats

PURPOSE: The aim of this study was to use laser Doppler flowmetry to measure anterior choroidal blood flow in the anesthetized rat and to determine the role of nitric oxide (NO) in the maintenance of basal ocular blood flow in vivo. METHODS: By using laser Doppler flowmetry, blood flow from the anterior choroid in pentobarbital-anesthetized rats was measured continuously. Graded single doses (0.03-300 mg/kg) of the nonselective NO synthase inhibitor NG-nitro-L-arginine-methyl ester (L-NAME) were administered intravenously to establish dose-response relationships. Other groups of animals were tested with L-NAME after the prior administration of L-arginine, with D-NAME, or with the selective neural NO synthase inhibitor 7-nitroindazole. RESULTS: Intravenous administration of L-NAME produced a dose-related depression of anterior choroidal blood flow in the 0.3- to 30-mg/kg range. Maximal depression of approximately 60% occurred at the 30-mg/kg dose, peaked at approximately 30 minutes, and lasted throughout the 60-minute experimental period. At 10 mg/kg, L-NAME reduced ocular blood flow by approximately 50%, an effect that was abolished by pretreatment with intravenous L-arginine (300 mg/kg). Both D-NAME (10 mg/kg, intravenously) and 7-nitroindazole (50 mg/kg, intraperitoneally) were inactive with regard to ocular blood flow depression. CONCLUSIONS: Laser Doppler flowmetry appears to be a useful tool for continuous, online measurement of anterior choroidal blood flow in the rat eye. Results with L-NAME and 7-nitroindazole suggest that local tonic generation of endothelial NO plays an important role in the maintenance of basal anterior choroidal blood flow in this species.