Biomedical subjects

E Hamel

Publications and source records attributed to E Hamel.

At least 127 records · Page 7Linked to original sources

PubMed1995

Pharmacological characterization of muscarinic acetylcholine binding sites in human and bovine cerebral microvessels.

The aim of this study was to investigate the presence of muscarinic receptors in human brain microvessels (MVs) and capillaries (CAPs) and, further, to pharmacologically characterize these receptors as well as those in bovine cerebral microvascular beds. For this purpose, the binding of [3H]N-methyl scopolamine ([3H]NMS) was assessed in isolated human and bovine cerebral MVs and CAPs and competition studies were performed against [3H]NMS binding with several well characterized muscarinic antagonists. The antagonist cerebrovascular affinity constants (pKD) were determined with the computer-fitting software LIGAND and then compared by correlation analyses to their reported affinities (pKi) at the five cloned muscarinic receptors. The specific binding of [3H]NMS to human and bovine MVs and CAPs was saturable, of high affinity and competitively inhibited by muscarinic antagonists. Heterogeneous populations of muscarinic binding sites were found in the microvascular tissues from both species. In human cortical MVs, the pharmacological binding profile obtained from various muscarinic receptor antagonists was best correlated to that of the cloned ml (r = 0.95; p < 0.001) and less so m5 (r = 0.77; p = 0.025) receptor subtypes while in bovine MVs, the presence of the m1 subtype was strongly suggested. Cerebrovascular affinities obtained for selected muscarinic antagonists in single preparations of human and bovine CAPs were suggestive of the presence of M1/m1 and M3/m3 receptor subtypes, and possibly the m5 subtype in bovine CAPs. The detection of M1/m1, M3/m3 and possibly m5 muscarinic receptor subtypes in brain microcirculation is consistent with reports where these receptors have been shown to mediate vasoconstriction, vasodilatation, and activation of nitric oxide synthase, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Aged↗

PubMed1995

Ultrastructural analysis of tryptophan hydroxylase immunoreactive nerve terminals in the rat cerebral cortex and hippocampus: their associations with local blood vessels.

Physiological evidence has indicated that serotonin (5-hydroxytryptamine) could be a regulator of cerebral blood flow in various regions of the brain. In the present study, tryptophan hydroxylase immunocytochemistry was used to characterize, both at the light and electron microscopic levels, serotonergic nerve terminals and primarily their relationships with intraparenchymal microarterioles and capillaries in the rat frontoparietal cortex, entorhinal cortex and hippocampus. Irrespective of the brain area, serotonergic varicosities were primarily apposed to either dendrites or nerve terminals, were on average 0.37 micron2 in surface area (0.69 micron calculated diameter) and 12-22% of them engaged in synaptic junctions, mostly with dendritic elements. Perivascular terminals (defined as immunolabelled varicosities located within a 3 micron perimeter around the vessel basal lamina) in the frontoparietal cortex represented 8-11% of all immunoreactive terminals counted, as determined by light and electron microscopy, respectively. In the entorhinal cortex and hippocampus, the proportion of perivascular terminals was only determined at the ultrastructural level and corresponded to 10% and 4%, respectively. In the frontoparietal cortex, serotonergic varicosities were located significantly closer (n = 250, 0.98 +/- 0.05 micron; P < 0.001) to the blood vessels than those of the entorhinal cortex (n = 116, 1.41 +/- 0.08 microns) or hippocampus (n = 105, 1.31 +/- 0.08 microns). Of all perivascular serotonergic terminals in the frontoparietal cortex, 26% were in the immediate vicinity (0-0.25 micron) of the vessel wall, with 2.8% directly abutting on the basement membrane, while 11.6% were separated from it only by a thin astrocytic leaflet. This situation contrasts with that observed in the entorhinal cortex and hippocampus, where no immunoreactive varicosity was ever seen directly contacting the vessel basal lamina and with only 10-13% of the terminals being within 0.25 micron from the vessels. The surface area of perivascular serotonergic terminals was comparable in all regions studied and corresponded to 0.22 micron2; these virtually never engaged in synaptic contacts with adjacent neuronal structures. Our results indicate that tryptophan hydroxylase-immunolabelled terminals are identical to previously characterized serotonin-containing varicosities. Furthermore, the present data show intimate associations between serotonergic terminals and microvessels in the three regions examined. However, perivascular terminals in the frontoparietal cortex were more frequent and/or located much closer to local microvessels than those in the other regions, and might be more directly involved in neurogenic control of local cerebral blood flow.

Animals↗

PubMed1995

Antitumor agents, 154. Cytotoxic and antimitotic flavonols from Polanisia dodecandra.

Three flavonols, 5,3'-dihydroxy-3,6,7,8,4'-pentamethoxyflavone [1], 5,4'-dihydroxy-3,6,7,8,3'-pentamethoxyflavone [2], and quercetin 3-O-beta-D-glucopyranosyl-7-O-alpha-L-rhamnopyranoside [3], were isolated from Polanisia dodecandra. Compound 1 showed remarkable cytotoxicity in vitro against panels of central nervous system cancer (SF-268, SF-539, SNB-75, U-251), non-small cell lung cancer (HOP-62, NCI-H266, NCI-H460, NCI-H522), small cell lung cancer (DMS-114), ovarian cancer (OVCAR-3, SK-OV-3), colon cancer (HCT-116), renal cancer (UO-31), a melanoma cell line (SK-MEL-5), and two leukemia cell lines (HL-60 [TB], SR), with GI50 values in the low micromolar to nanomolar concentration range. This substance also inhibited rubulin polymerization (IC50 = 0.83 +/- 0.2 microM) and the binding of radiolabeled colchicine to tubulin with 59% inhibition when present in equimolar concentrations with colchicine. Compound 2 also showed cytotoxicity against medulloblastoma (TE-671) tumor cells with an ED50 value of 0.98 microgram/ml. Compound 1 appears to be the first example of a flavonol to exhibit potent inhibition of tubulin polymerization and, therefore, warrants further investigation as an antimitotic agent.

Antineoplastic Agents↗

PubMed1995

Characterization of neuropeptide Y (NPY) receptors in human cerebral arteries with selective agonists and the new Y1 antagonist BIBP 3226.

1. We have characterized pharmacologically the receptor subtype(s) responsible for the neuropeptide Y (NPY)-induced vasoconstriction in human cerebral arteries. NPY, PYY and several of their derivatives with well defined affinities at the known Y1 and Y2 receptor subtypes were used. Moreover, we tested the ability of the new Y1 receptor antagonist, BIBP 3226, to antagonize the NPY-induced cerebral vasoconstriction. 2. NPY, PYY and their agonists with high affinities at the Y1 receptor subtype ([Leu31-Pro34]-NPY and [Leu31-Pro34]-PYY) elicited strong, long lasting and concentration-dependent contractions of human cerebral arteries. Compounds with Y2 affinity such as PYY3-36 or NPY13-36 either elicited a submaximal contraction at high concentrations or failed to induce any significant vasomotor response. Also, the application of NPY or the specific Y1 agonist, [Leu31-Pro34]-NPY, to human cerebral vessels pretreated with the Y1 agonist, NPY13-36, resulted in contractile responses identical to those obtained when these compounds were tested without prior application of NPY13-36. 3. The order of agonist potency at the human cerebrovascular receptor was: [Leu31-Pro34]-NPY = [Leu31-Pro34]-PYY > or = NPY > PYY > PYY3-36 > > > NPY13-36, which corresponded to that reported previously at the neuronal and vascular Y1 receptors. 4. Increasing concentrations (10(-9)-10(-6) M) of the Y1 receptor antagonist, BIBP 3226, to human cerebral vessels caused a parallel and rightward shift in the NPY dose-response curves without any significant change in the maximal contractile response. The calculated pA2 was 8.52 +/- 0.13, a value compatible with the reported affinity at the rodent and human Y1 receptor. 5. We conclude that Y1 receptors exclusively, mediate the NPY-induced contraction in human cerebral arteries and we show that BIBP 3226 is a potent and competitive antagonist of this YI-mediated vasoconstriction.

Adult↗

PubMed1995

Cholinergic basal forebrain neurons project to cortical microvessels in the rat: electron microscopic study with anterogradely transported Phaseolus vulgaris leucoagglutinin and choline acetyltransferase immunocytochemistry.

Physiological evidence indicates that central cholinergic pathways are involved in the regulation of cerebral cortical blood flow. We investigated the possible contribution of basal forebrain cholinergic neurons from the substantia innominata (SI) to the innervation of cortical microvessels. Basalo-cortical perivascular nerve terminals were detected by light and electron microscopic immunocytochemistry of anterogradely transported Phaseolus vulgaris leucoagglutinin (PHA-L) following its injection in the SI, and were compared to cortical perivascular cholinergic (immunoreactive for choline acetyltransferase (ChAT)) terminals. The basal forebrain origin of cholinergic terminals was evaluated after unilateral ibotenic acid lesion of the SI. PHA-L varicose fibers reached and surrounded microvessels in all cortical subdivisions examined. When studied at the ultrastructural level in the fronto-parietal and perirhinal cortices, perivascular PHA-L nerve terminals were located significantly closer to microvessels in the fronto-parietal than in the perirhinal cortex (respective average distance of 0.98 +/- 0.09 and 1.34 +/- 0.07 micron; p < 0.01). PHA-L and ChAT terminals in the fronto-parietal cortex compared very well in their perivascular distribution but not in the perirhinal cortex. In both cortices, perivascular PHA-L terminals were similar in size to, but engaged more frequently in synaptic contacts than their ChAT counterparts. Following SI lesion, the density of cortical ChAT terminals including those reaching microvessels decreased significantly (56 and 63%, respectively, p < 0.005) in the fronto-parietal cortex, while the cortical and perivascular denervations were much less pronounced (26%, not significant and 35%, p < 0.05, respectively) in the perirhinal cortex. These results indicate that basal forebrain neurons project preferentially to fronto-parietal cortical microvessels and further show that a significant proportion of these projections are cholinergic. In addition, the difference in distribution and/or synaptic incidence between perivascular PHA-L and ChAT terminals suggested that noncholinergic SI neurons also contribute to these neurovascular associations, and more so in the perirhinal cortex, as further indicated by the lesion studies. Such finding corroborates recent physiological evidence for a functional innervation of the cortical microvascular bed by SI neurons, a role that might be relevant to the overall pathology of Alzheimer's dementia.

Animals↗

PubMed1995

Antineoplastic agents 337. Synthesis of dolastatin 10 structural modifications.

New structural modifications of the marine shell-less mollusk peptide constituent dolastatin 10 (1) have been synthesized, and evaluated against a variety of cancer cell lines and for their ability to inhibit tubulin polymerization. A number of useful structure-activity relationships were uncovered. The most important observation was that the dolaphenine unit of dolastatin 10 could be satisfactorily replaced with a phenethylamine. Peptide 11C, designated auristatin PE, was found to exhibit inhibition of cancer cell growth and tubulin assembly comparable to that of dolastatin 10.

Amino Acid Sequence↗

PubMed1995

Characterization of the interaction of the marine cyanobacterial natural product curacin A with the colchicine site of tubulin and initial structure-activity studies with analogues.

Curacin A, the major lipid constituent of a strain of the marine cyanobacterium Lyngbya majuscula obtained off the coast of Curaçao, is a potent antimitotic agent that we have previously shown to inhibit microtubule assembly and colchicine binding to tubulin. In the present study, we report that curacin A probably binds in the colchicine site because it competitively inhibits the binding of [3H]colchicine to tubulin with an apparent Ki value of 0.6 microM and stimulates tubulin-dependent GTP hydrolysis, as do most other colchicine-site agents. The binding of curacin A to tubulin resembled the binding reactions of combretastatin A-4 and podophyllotoxin in contrast to that of colchicine in that it occurred as extensively on ice as at higher temperatures. However, once bound, the dissociation rate of curacin A from tubulin is very slow, more closely resembling that observed with colchicinoids (thiocolchicine was the drug examined) than the faster dissociation that occurs with combretastatin A-4 and podophyllotoxin. Because the molecular structure of curacin A is so different from that of previously described colchicine-site drugs (e.g., there is no aromatic moiety, and there are only two conjugated double bonds in its linear hydrocarbon chain), we have been examining the activities of natural isomers and synthetic derivatives. So far, only modest enhancement or reduction of activity has been observed with a variety of structural changes.

Animals↗

PubMed1995

Interaction of dolastatin 10 with tubulin: induction of aggregation and binding and dissociation reactions.

We have prepared [3H]dolastatin 10 and examined its interactions with tubulin. Binding kinetics appeared to be biphasic, with a rapid initial reaction that could not be accurately measured, followed by a slower second reaction. Bound drug was stable in centrifugal gel filtration, column gel filtration, and high performance liquid chromatography gel filtration, but the bound drug could be displaced by an active isomer of dolastatin 10. Scatchard analysis of binding data was consistent with two classes of binding sites. However, dolastatin 10 induced an aggregation reaction upon binding to tubulin, complicating analysis of the data, and incorporation of [3H]dolastatin 10 into large aggregates was readily demonstrated. The chromatographic properties of the smallest radiolabeled species that could be documented were most consistent with a complex consisting of two molecules of alpha/beta-tubulin dimer and two molecules of [3H]dolastatin 10. The coexistence of an aggregation reaction with a binding reaction at a single site probably underlies the biphasic binding kinetics and the biphasic Scatchard plot. Of peptides that strongly inhibit tubulin polymerization (dolastatin 10, dolastatin 10 isomers, segments, and analogs, dolastatin 15, and phomopsin A), only those previously shown to be strong inhibitors of vinblastine binding and nucleotide exchange also strongly inhibited [3H]dolastatin 10 binding and induced tubulin aggregation (dolastatin 10 itself, two chiral isomers of dolastatin 10, and phomopsin A). The morphology of dolastatin 10-induced aggregates was compared with that of vinblastine-induced aggregates under a variety of reaction conditions. With both drugs the aggregates had a more organized appearance when microtubule-associated proteins were included in the reaction.

Animals↗

PubMed1994

Antitumor agents. 155. Synthesis and biological evaluation of 3',6,7-substituted 2-phenyl-4-quinolones as antimicrotubule agents.

A series of 3',6,7-substituted 2-phenyl-4-quinolones were designed and synthesized as antimitotic antitumor agents. All compounds showed cytotoxic effects (log GI50 < or = -4.0; log drug molar concentration required to cause 50% inhibition) against the growth of a variety of human tumor cell lines, including those derived from solid tumors such as non-small cell lung, colon, central nervous system, ovary, prostate, and breast cancers, when evaluated in the National Cancer Institute's 60 human tumor cell line in vitro screen. The most potent compound (26) demonstrated strong cytotoxic effects with GI50 values in the nanomolar or subnanomolar range in almost all the tumor cell lines. Compound 26 was also a potent inhibitor of tubulin polymerization and radiolabeled colchicine binding to tubulin, with activity comparable to those of the potent antimitotic natural products colchicine, podophyllotoxin, and combretastatin A-4.

Animals↗

PubMed1994

Distinct choline acetyltransferase (ChAT) and vasoactive intestinal polypeptide (VIP) bipolar neurons project to local blood vessels in the rat cerebral cortex.

Innervation of rat intracortical cerebral blood vessels by acetylcholine (ACh) and vasoactive intestinal polypeptide (VIP) remains largely unexplored and it is not known if the cells of origin are intra- or extracortical nor if perivascular fibers colocalize ACh and VIP. Cortical cholinergic innervation arises primarily from the basal forebrain and to a small extent from intrinsic bipolar ACh neurons thought to be the sole source of cortical VIP. In order to evaluate if intracortical perivascular ACh terminals could be distinguished from those of the basal forebrain by their colocalization with VIP, we performed a double immunofluorescence study and determined the percentage of colocalization of choline acetyltransferase (ChAT) and VIP in cortical neurons, as well as in terminal fields associated with intracortical blood vessels. From a total of 2103 cells examined in all cortical areas, VIP neurons accounted for the largest population (58.3%) followed by ChAT-positive cells (28.2%) with only 13.5% of cells being double-labelled for VIP and ChAT. Of the cortical ChAT-immunostained cells (n = 878), 32.3% colocalized VIP whereas only 18.8% of VIP neurons (n = 1509) also contained ChAT. In various cortical areas, ChAT cell bodies were seen to be contacted by VIP terminals which surrounded closely their cell soma and proximal dendrites. Perivascular fibers studied by double immunofluorescence and confocal microscopy were of three categories including cholinergic, VIPergic with a smaller population of fibers which costained for both ChAT and VIP. These results show that cortical VIP neurons are much more numerous than those containing ChAT, and that a majority of VIP neurons do not colocalize with ChAT. This observation indicates that ACh and VIP are primarily located in distinct neuronal populations and that VIP cannot be used as a marker of intracortical ACh neurons and terminals. Our results further suggest that intracortical blood vessels are primarily under the influence of distinct ChAT and VIP perivascular fibers. Also, the presence of a subset of VIP and ChAT/VIP fibers in association with intracortical blood vessels strongly suggests a role for cortical bipolar neurons in local cerebrovascular regulation. The origin of the perivascular ChAT fibers which do not colocalize VIP, however, remains unknown.

Animals↗

PubMed1994

The magnesium-GTP interaction in microtubule assembly.

Microtubule-associated-protein-dependent assembly of tubulin with GDP in the exchangeable site (tubulin-GDP) can occur with minimal free Mg2+ (< 3 microM). This reaction is totally inhibited by EDTA and by GTP concentrations over 2 mM and stimulated by MgCl2. Quantitative aspects of this stimulation are affected by both the Mg2+ and GTP concentrations but no relationship exists between reaction rates and relative amounts of different magnesium and GTP species. GTP binding to tubulin-GDP, while maximally stimulated 2-3-fold by exogenous MgCl2, was inhibited less than 50% by EDTA, and the amount of GTP bound increased as its concentration rose to levels that inhibited polymerization. Studies on the binding of Mg2+ to tubulin-GDP in the presence and absence of GTP showed that the increase in the amount of tubulin-associated Mg2+ was substoichiometric to the amount of GTP bound (maximum stoichiometry of additional Mg2+ to GTP bound, 0.7). Upon polymerization the increased Mg2+ content of tubulin was reduced, indicating its loss during GTP hydrolysis. Mg2+ thus plays a critical role in assembly distinct from its enhancement of GTP binding to the exchangeable site. If magnesium is present in trace amounts, this role must either be catalytic during polymerization or limited to nucleation.

Animals↗

PubMed1994

Light and electron microscopic immunocytochemical analysis of the neurovascular relationships of choline acetyltransferase and vasoactive intestinal polypeptide nerve terminals in the rat cerebral cortex.

Acetylcholine or vasoactive intestinal peptide (VIP) nerve terminals closely related to intracortical blood vessels have previously been reported. Recent physiological evidence indicates that these central neuronal systems are involved in the fine control of local cerebral blood flow. In the present study, the intimate associations between choline acetyltransferase (ChAT) and VIP axon terminals and intracortical microvessels were characterized by light (LM) and electron microscopic (EM) immunocytochemistry. In semithin sections, LM analysis of the distribution of ChAT- and VIP-immunostained puncta juxtaposed to small intraparenchymal blood vessels demonstrated that neither type of terminal was enriched or impoverished around microvessels within the cerebral cortex. At the EM level, most ChAT- or VIP-immunolabelled elements located within a 3 microns perimeter around vessel walls were axon terminals. These perivascular terminals were associated primarily with capillaries but also, to a lesser extent, with microarterioles. Even though ChAT and VIP terminals were frequently found in the immediate vicinity (< or = 0.25 microns) of microvessels, they almost never contacted the outer basal lamina, usually abutting onto perivascular astroglial leaflets. There were no membrane specializations at the site of contact between ChAT or VIP terminals and perivascular astroglia. In all cortical areas examined, the average size of VIP-immunolabelled varicosities (0.56 +/- 0.04 microns 2) was significantly larger than that of their ChAT counterparts (0.32 +/- 0.02 microns 2; P < 0.001). Perivascular VIP terminals were more frequently engaged in synaptic contact than those immunostained for ChAT, which rarely exhibited a synaptic junction even in serial thin sections. Neither VIP nor ChAT immunostaining was ever observed in endothelial cells. These results suggest that both acetylcholine and VIP exert their effects on intracortical microvessels through indirect, paracrine mechanisms. The marked difference in synaptic incidence and average size between both types of perivascular terminals indicates that these two vasoactive agents are primarily located in distinct neuronal populations. Further, our results show that the astrocytic glia is the major direct target for both ChAT and VIP perivascular terminals and suggest that neuronal/glial/vascular interactions are a key element in the neurogenic control of the intracortical microcirculation.

Animals↗

PubMed1994

2-Methoxyestradiol, an endogenous mammalian metabolite, inhibits tubulin polymerization by interacting at the colchicine site.

A metabolite of estradiol, 2-methoxyestradiol (2ME), inhibits angiogenesis in the chicken embryo chorioallantoic membrane assay. Since 2ME causes mitotic perturbations, we examined its interactions with tubulin. In our standard 1.0 M glutamate system (plus 1.0 mM MgCl2 at 37 degrees C), superstoichiometric concentrations (relative to tubulin) of 2ME inhibited the nucleation and propagation phases of tubulin assembly but did not affect the reaction extent. Although polymer formed in the presence of 2ME was more cold-stable than control polymer, morphology was little changed. Under suboptimal reaction conditions (0.8 M glutamate/no MgCl2 at 26 degrees C), substoichiometric 2ME totally inhibited polymerization. No other estrogenic compound was as effective as 2ME as an inhibitor of polymerization or of the binding of colchicine to tubulin. Inhibition of colchicine binding was competitive (Ki, 22 microM). Thus, a mammalian metabolite of estradiol binds to the colchicine site of tubulin and, depending on reaction conditions, either inhibits assembly or seems to be incorporated into a polymer with altered stability properties.

2-Methoxyestradiol↗

PubMed1994

Antitumor agents. 150. 2',3',4',5',5,6,7-substituted 2-phenyl-4-quinolones and related compounds: their synthesis, cytotoxicity, and inhibition of tubulin polymerization.

As part of our continuing search for potential anticancer drug candidates in the 2-phenyl-4-quinolone series, we have synthesized a series of 6,7-methylenedioxy-substituted and unsubstituted 2-phenyl-4-quinolones, as well as related compounds. Their in vitro inhibition of human tumor cell lines and tubulin polymerization is reported. In general, a good correlation was found between cytotoxicity and inhibition of tubulin polymerization. Compounds 7, 9, 13, 16, 22, 23, 36, and 37 showed potent inhibitory effects in both assays. All rigid analogs (47-49) and trimethoxy-substituted compounds showed little or no activity. Substitution at the 4'-position also resulted in compounds with little or no activity, except for hydroxyl or methyl groups at this position. Further investigation is underway to determine if substitution at the 3'-position will result in compounds with increased activity.

Animals↗

PubMed1994

Cholinergic dilatation and constriction of feline cerebral blood vessels are mediated by stimulation of phosphoinositide metabolism via two different muscarinic receptor subtypes.

The muscarinic receptors involved in phosphoinositide (PI) hydrolysis have been pharmacologically characterized in cat cerebral blood vessels. Carbachol elicited a concentration-dependent increase in inositol phosphate accumulation [inositol monophosphate, bisphosphate, trisphosphate (IP3) and tetrakisphosphate] in both major cerebral arteries and small pial vessels, which reached 140-280% of baseline at 10(-3) M carbachol (referred to as maximal effect). However, the inositol phosphate accumulation response was found to be biphasic with a submaximal effect (30-50% of the maximal stimulation) obtained at low carbachol concentrations (< 10(-5) M). Endothelial denudation induced a virtual disappearance of the submaximal PI response without affecting that elicited by high concentrations of carbachol. The pharmacology of the two carbachol-induced PI responses was investigated by comparing the potency of selected muscarinic antagonists to block the IP3 accumulation induced by 10(-7) M (endothelium-dependent submaximal effect) and 10(-4) M (endothelium-independent near-maximal effect) carbachol. In both major arteries and pial vessels, the activation of IP3 production by 10(-4) M carbachol was similarly inhibited by muscarinic antagonists with the following averaged rank order of potency (in -log IC50): 4-diphenylacetoxy-N-methylpiperidine methiodide (4-DAMP; 8.65) > pirenzepine (8.28) > 6-chloro-5,10-dihydro-5-[(1-methyl-4-piperidinyl)acetyl]-11H- dibenzo[b,e][1,4]diazepine-11-one (UH-AH 371; 7.87) > 11-[[2-[(diethylamino)methyl]-1-piperidinyl]acetyl]-5,-11- dihydro-6H-pyridol[2,3-b][1,4]benzodiazepine-6-one (AF-DX 116; 6.62), a pharmacological profile compatible with an M1 receptor subtype.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1993

Recovery of choline acetyltransferase activity without sprouting of the residual acetylcholine innervation in adult rat cerebral cortex after lesion of the nucleus basalis.

In view of the divergent literature concerning the long-term effects of ibotenic acid lesions of the nucleus basalis of Meynert (NBM) on the choline acetyltransferase (ChAT) activity in adult rat cerebral cortex, we have critically reassessed the issue of an eventual recovery of this enzymatic activity by sprouting of the residual acetylcholine (ACh) innervation. At short (1 week) and long survival time (3 months) after unilateral ibotenic acid lesion, ChAT activity was biochemically measured in the ipsi and contralateral fronto-parietal cortex of several rats in which the extent of ACh neuronal loss in NBM was also estimated by counts of ChAT-immunostained cell bodies on the lesioned vs. non-lesioned side. In other lesioned rats, particular attention was paid to the distribution of the residual cortical ACh (ChAT-immunostained) innervation, and that of immunostained vasoactive intestinal polypeptide (VIP) axon terminals known to belong in part to intrinsic cortical ACh neurons which co-localize this peptide. One week after NBM lesion, profound decreases of ipsilateral cortical ChAT activity were tightly correlated with the extent of ACh cell body loss in the nucleus. A significant recovery of cortical ChAT activity could be documented after 3 months, despite persistence of NBM cell body losses as severe as after 1 week. At both survival times, the number of ChAT-immunostained axons was markedly reduced throughout the ipsilateral fronto-parietal cortex, demonstrating that most ACh fibers of extrinsic origin had been permanently removed. This result also indicated that the long-term recovery of ChAT activity had occurred without sprouting of the residual ACh innervation. The laminar distribution and number of VIP-immunostained terminals remained the same on the lesioned and intact side and comparable to normal, ruling out an extensive sprouting of intrinsic ACh/VIP or VIP alone fibers. The return to a near normal cortical ChAT activity in severely ACh-denervated cortex suggested that the intrinsic ACh innervation was primarily responsible for this recovery.

Acetylcholine↗

PubMed1993

5-HT1 receptors mediating contraction in bovine cerebral arteries: a model for human cerebrovascular '5-HT1D beta' receptors.

We report on the pharmacological profile of the 5-HT receptor which induces contraction of the bovine isolated cerebral arteries. Several 5-HT receptor agonists were tested for their ability to induce vasoconstriction in bovine pial arteries and their potencies were compared to that of 5-HT. The rank order of agonist potency can be summarized as 5-carboxamidotryptamine (5-CT) = RU 24969 > or = 5-HT > sumatriptan > alpha-methyl-5-HT > methysergide > 2-methyl-5-HT > ((+-)-2-dipropylamino-8-hydroxy-1,2,3,4-tetrahydronaphthalene (8-OH-DPAT). Only methysergide induced a contraction which was smaller than that elicited by 5-HT. Antagonists with selective affinity at 5-HT1A/1B (propranolol), 5-HT1C (mesulergine), 5-HT2 (ketanserin, mianserin) and 5-HT3 (MDL 72222) sites were inactive to block the 5-HT-induced contraction. In contrast, the 5-HT1/5-HT2 receptor antagonists methiothepin and metergoline inhibited the 5-HT-induced response with relatively high affinity (pA2 = 8.16 +/- 0.26 and 6.73 +/- 0.05, respectively). Overall, this pharmacological profile indicated clearly that a 5-HT1 receptor, most closely related to the 5-HT1D subtype, is responsible for the 5-HT-induced contraction of bovine cerebral arteries. Correlation analysis of the potencies of a series of 5-HT receptor agonists and antagonists in bovine and human cerebrovascular preparations showed a highly significant positive correlation (r = 0.94, P = 0.0051).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1993

Synthesis and evaluation of a series of benzylaniline hydrochlorides as potential cytotoxic and antimitotic agents acting by inhibition of tubulin polymerization.

Although certain substituted cis-stilbenes have displayed potent tubulin polymerization inhibitory activity and significant cytotoxicities in cancer cell cultures, these compounds have limited aqueous solubility and are therefore difficult to formulate for in vivo evaluation. A series of water-soluble N-(3,4,5-trimethoxybenzyl)aniline salts has therefore been synthesized in which the olefinic bridge of the stilbenes is replaced by an aminomethylene hydrochloride moiety. A relationship was found between the size of the substituent in the 4-position of the aniline ring and both antitubulin activity and cytotoxicity, such that the smaller the substituent, the greater the potency. The most promising of the newly synthesized compounds was 4-methyl-N-(3,4,5-trimethoxybenzyl)aniline hydrochloride, with an IC50 value of 3.5 microM for inhibition of tubulin polymerization and cytotoxicity for a wide variety of cancer cell lines. The cytotoxicities of the benzylaniline hydrochlorides correlated remarkably well with their antitubulin activities.

Aniline Compounds↗