Biomedical subjects

M Gola

Publications and source records attributed to M Gola.

At least 37 records · Page 2Linked to original sources

PubMed1999

Pharmacological properties of trimebutine and N-monodesmethyltrimebutine.

Trimebutine [2-dimethylamino-2-phenylbutyl-3,4,5-trimethoxybenzoate hydrogen maleate (TMB)] has been demonstrated to be active for relieving abdominal pain in humans. To better understand its mechanism of action, we have tested TMB; nor-TMB, its main metabolite in humans; and their respective stereoisomers for their affinity toward sodium channels labeled by [3H]batrachotoxin, their effect on sodium, potassium, and calcium currents in rat dorsal root ganglia neurons, and their effect on veratridine-induced glutamate release from rat spinal cord slices. TMB has also been tested in an animal model of local anesthesia. TMB (Ki = 2.66 +/- 0.15 microM) and nor-TMB (Ki = 0.73 +/- 0.02 microM) displaced [3H]batrachotoxin from its binding site with affinities similar to that of bupivacaine (Ki = 7.1 +/- 0.9 microM). nor-TMB was found to block veratridine-induced glutamate release with an IC50 value of 8.5 microM, which is very similar to that of bupivacaine (IC50 = 8.2 microM); the effect of TMB was limited to 50% inhibition at 100 microM. TMB and nor-TMB blocked sodium currents in sensory neurons from rat dorsal root ganglia (IC50 = 0.83 +/- 0.09 and 1.23 +/- 0.19 microM, respectively), whereas no effect was observed on calcium currents at the same concentrations. A limited effect was observed on potassium currents (IC50 = 23 +/- 6 at 10 microM) for TMB. In vivo, when tested in the rabbit corneal reflex, TMB displayed a local anesthetic activity 17-fold more potent than that of lidocaine.

Animals↗

PubMed1999

Distribution in rat brain of binding sites of kaliotoxin, a blocker of Kv1.1 and Kv1.3 alpha-subunits.

The distribution of the binding sites for kaliotoxin (KTX), a blocker of voltage-dependent K(+) channels, was studied with quantitative autoradiography in adult rat brain and during postnatal brain maturation. Iodinated KTX bound specifically to tissue sections with a high affinity (K(d) = 82 pM) and a maximal binding capacity of 13.4 fmol/mg protein. The distribution of KTX binding sites within the central nervous system was heterogeneous. The highest densities were found in the neocortex, hypothalamus, dentate gyrus, bed nucleus of the stria terminalis, and parabrachial nuclei. The lowest level was observed in the white matter. From postnatal day 5 onward, KTX binding sites were detectable only in the hindbrain. The density of KTX binding sites in whole brain drastically increased after postnatal day 15 to achieve adult levels at postnatal day 60 in the whole brain. Bath application of KTX to Xenopus laevis oocytes blocked recombinant Kv1.3 and Kv1.1 channels potently and Kv1.2 channels less potently, with respective K(d) values of 0.1, 1.5, and 25 nM. KTX affinities for each of these channels expressed in mammalian cells were about 10-fold lower. A comparison of the distribution of KTX binding sites with that of Kv1 channel polypeptides, together with the pharmacology of KTX block, suggests that the principal targets for KTX in rat brain are K(+) channels containing Kv1.1 and Kv1.3 alpha-subunits.

Aging↗

PubMed1998

Encoding properties induced by a persistent voltage-gated muscarinic sodium current in rabbit sympathetic neurones.

1. A time- and voltage-dependent Na(+)-selective current termed INa,M is activated by muscarinic agonists or splanchnic nerve stimulation in sympathetic neurones of rabbit coeliac and superior mesenteric ganglia. The firing patterns induced by INa,M were investigated in patch-clamped neurones within intact ganglia, and compared with those generated by a neuronal model including INa,M. 2. INa,M was characterized by voltage-dependent low threshold activation and high-threshold inactivation functions. The overlapping functions produced a persistent U-shaped current between -100 and -20 mV, which peaked at the cell resting potential. The activation and inactivation kinetics were fitted to single exponentials with time constants of approximately 100 and 400 ms, respectively. 3. Activating INa,M with muscarinic agonists or nerve stimulation depolarized and fired the neurones. The depolarization was paralleled by an apparent increase in input membrane resistance. The model showed that this paradox resulted from the turning off of INa,M during resistance tests, which also accounted for the all-or-none slow hyperpolarizing responses to current pulses. 4. INa,M gave the neurones an N-shaped I-V relationship capable of producing complex firing patterns. Under given conditions, carbachol-treated neurones could either fire regularly or remain silent at approximately -80 mV, i.e. they displayed bistability. Transitions from one state to the other were triggered with short current pulses. The transitions resulted from the turning on and off of INa,M. 5. Firing reduced INa,M, an effect abolished by blocking Ca2+ channels or adding BAPTA (40 mM) to the pipette. The Ca(2+)-related negative regulation of INa,M may have mediated endogenous bursting activity. Burst firing was generated by the model upon introducing Ca2+ regulation of INa,M. 6. The results demonstrate that INa,M gives prevertebral sympathetic neurones a wide repertoire of firing patterns: pacemaker-like properties, bistability and burst firing capability. They suggest that the INa,M-related encoding properties may provide sympathetic neurotransmission with new potentialities.

Animals↗

PubMed1998

Structural basis for specificity and potency of xanthine derivatives as activators of the CFTR chloride channel.

1. On the basis of their structure, we compared the ability of 35 xanthine derivatives to activate the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel stably expressed in chinese hamster ovary (CHO) cells using the cell-attached patch clamp and iodide efflux techniques. 2. Activation of CFTR channels was obtained with 3-mono, 1,3-di or 1,3,7-tri-substituted alkyl xanthine derivatives (enprofylline, theophylline, aminophylline, IBMX, DPMX and pentoxifylline). By contrast, xanthine derivatives substituted at the C8- or N9-position failed to open CFTR channels. 3. The CFTR chloride channel activity was blocked by glibenclamide (100 microM) but not by DIDS (100 microM). 4. Activation of CFTR by xanthines was not mimicked by the calcium ionophore A23187, adenosine, UTP, ATP or the specific phosphodiesterase inhibitors rolipram, Ro 20-1724 and milrinone. In addition, we found no correlation between the effect of xanthines on CFTR and on the cellular cyclic AMP or ATP levels. 5. We then synthesized a series of 3,7-dimethyl-1-alkyl xanthine derivatives; among them, 3,7-dimethyl-1-propyl xanthine and 3,7-dimethyl-1-isobutyl xanthine both activated CFTR channels without increasing the intracellular cyclic AMP level, while the structurally related 3,7-dimethyl-1-(2-propenyl) xanthine and 3,7-dimethyl-1-(oxiranyl methyl) xanthine were inactive. 6. Our findings delineate a novel function for xanthine compounds and identify the molecular features that enable xanthine activation of CFTR. These results may be useful in the development of new molecules for studying the pharmacology of chloride channels.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

PubMed1998

[Efficacy of delayed-release lanreotide in the medical therapy of acromegaly].

BACKGROUND: The efficacy of 6 months therapy with slow-release lanreotide (30 mg i.m. every 10-14 days) in 8 acromegalic patients has been studied. METHODS: These patients had been previously treated (for 62 +/- 5.7 months) with octreotide (100 micrograms t.i.d.) and therefore presented, at the beginning of the study, normal mean GH (3.5 +/- 1.1 ng/ml) and IGF-1 (301.7 +/- 32.9 ng/ml) plasma levels. After a week of wash-out, mean GH (5.5 +/- 1.3 ng/ml) and IGF-1 (523.8 +/- 26.7 ng/ml) plasma levels showed a significant increase (p < 0.01) compared to the values observed during the treatment with octreotide. All 8 acromegalic patients then started the treatment with lanreotide, 30 mg i.m. After 14 days, mean GH plasma levels (4.2 +/- 1.3 ng/ml) did not significantly differ (p = NS) from those observed in the same group of patients during treatment with octreotide, whilst plasma IGF-1 levels (477 +/- 43 ng/ml) were significantly higher (p < 0.05). Four patients, in which mean plasma GH values resulted < 5 ng/ml, continued the therapy with lanreotide every 14 days. In the remaining 4 patients, in which plasma GH values were > 5 ng/ml, lanreotide was administered every 10 days. RESULTS: After 3 months of therapy, 6 out of the 8 patients presented persistent GH levels < 5 ng/ml during the day, with IGF-1 levels comparable to those observed during treatment with octreotide. The other 2 subjects presented plasma GH levels > 5 ng/ml during the day, with increased plasma levels of IGF-1. This latter group of patients was resubmitted treatment with octreotide, 100 micrograms t.i.d. After 6 months of therapy, all 6 patients presented GH and IGF-1 plasma levels comparable to those observed during treatment with octreotide. CONCLUSIONS: These data show that slow-release lanreotide can be a valid therapeutic alternative to octreotide in the medical treatment of acromegalic patients.

Acromegaly↗

PubMed1997

A potassium-channel toxin from the sea anemone Bunodosoma granulifera, an inhibitor for Kv1 channels. Revision of the amino acid sequence, disulfide-bridge assignment, chemical synthesis, and biological activity.

The potassium channel toxin secreted by the sea anemone Bunodosoma granulifera (BgK) is a 37-amino-acid peptide containing three disulfide bridges. Because a synthetic peptide corresponding to the reported sequence of BgK was found not to fold properly, the sequence was determined again. The new sequence differed from the previous one in the C-terminal tetrapeptide, which contains two cysteines involved in disulfide bridging. The revised sequence is: V C R D W F K E T A C R H A K S L G N C R T S Q K Y R A N C A K T C E L C. The toxin BgK was synthesized according to the new sequence and folded successfully. Disulfide bridges were assigned by peptide mapping on both natural and synthetic forms to be between Cys2-Cys37, Cys11-Cys30 and Cys20-Cys34. The toxin contains a C-terminal free carboxylate as shown by comparing the native toxin with two synthetic peptides containing the C-terminus in either the carboxylate or carboxamido form. Synthetic BgK inhibits binding of 125I-alpha-dendrotoxin to rat brain synaptosomal membranes, similarly to natural BgK (nanomolar range). No activity was observed on maxi-K+ channels incorporated into planar lipid bilayers. The ability of BgK to block voltage-dependent K+ channels was determined from recordings of whole cell currents in Xenopus oocytes injected with cRNA encoding three cloned Kv1 channels (Kv1.1, Kv1.2, Kv1.3) and one Kv3 (Kv3.1) channel. The Shaker-related Kv1 channels are equally affected by BgK, while the Shaw-related channel Kv3.1 is insensitive up to 0.125 microM toxin. Indeed, half blockage of the current through the three Kv1 channels tested occurred in the same concentration range (Kd = 6 nM for Kv1.1, 15 nM for Kv1.2, 10 nM for Kv1.3). The specificity of BgK for the Shaker-related K+ channels indicates that BgK is able to discriminate a large group of neuronal Kv1 channels in situ. The sequence, the disulfide bridge pattern, the secondary structure and the biological activity of BgK demonstrated that the sea anemone toxins, i.e. BgK, ShK and Kaliseptine, constitute novel molecular probes useful for investigating K+ channel properties.

Amino Acid Sequence↗

PubMed1997

Exotoxin-insensitive G proteins mediate synaptically evoked muscarinic sodium current in rabbit sympathetic neurones.

1. The involvement of G proteins in the transduction pathway that links muscarinic receptors to the low-threshold voltage-dependent sodium current (INa,M) was studied in neurones from intact sympathetic prevertebral ganglia using the whole-cell configuration of the patch-clamp technique. Experiments were performed in the presence of the nicotinic receptor antagonists hexamethonium (50 microM) and d-tubocurarine (50 microM). 2. INa,M was activated by either bath-applying muscarinic agonists or stimulating the preganglionic splanchnic nerves. Synaptically and agonist-mediated INa,M did not display significant run-down or changes in their properties in cells tested, irrespective of whether the pipette solutions contained GTP. 3. Dialysis of sympathetic neurones with GDP beta S (500-750 microM) decreased the amplitude of INa,M by approximately 65% compared with control neurones within 30 min. 4. In the absence of muscarinic receptor stimulation, intracellular dialysis with GTP gamma S (500 microM) for 10 min slowly and slightly (20-25%) activated INa,M. GTP gamma S dialysis markedly slowed down the decay of INa,M after its transient activation with carbachol pulses (10-20 s) or nerve stimulation (3-5 s). The INa,M activation became fully irreversible 2.9 min after the start of GTP gamma S dialysis. Dialysing cells with the G protein activator AIF4-led to a rapid but transient activation of INa,M. 5. Synaptically and agonist-evoked INa,M were not affected in ganglia treated with 0.5-1 microgram ml-1 pertussis toxin (PTX) for 7-24 h at 37 degrees C. Control experiments showed that this treatment severely reduced the PTX-sensitive inhibition of N-type calcium currents induced by carbachol (CCh) and noradrenaline. Application of NEM (N-ethylmaleimide) for 2 min depressed the INa,M evoked in response to bath-applied CCh by only 27%. 6. Incubating ganglia with 5-10 micrograms ml-1 of cholera toxin for 7 h had no effect on the carbachol-induced INa,M but greatly potentiated (approximately 250%) the synaptically evoked INa,M, presumably via a presynaptic mechanism. 7. These results show that the coupling between muscarinic receptors and NaM channels is mediated by pertussis toxin- and cholera toxin-insensitive G proteins, possibly of the Gq/11 or G12 class.

Animals↗

PubMed1997

ABC1, an ATP binding cassette transporter required for phagocytosis of apoptotic cells, generates a regulated anion flux after expression in Xenopus laevis oocytes.

The ATP binding cassette transporter ABC1 is a 220-kDa glycoprotein expressed by macrophages and required for engulfment of cells undergoing programmed cell death. Since members of this family of proteins such as P-glycoprotein and cystic fibrosis transmembrane conductance regulator share the ability to transport anions, we have investigated the transport capability of ABC1 expressed in Xenopus oocytes using iodide efflux and voltage-clamp techniques. We report here that ABC1 generates an anion flux sensitive to glibenclamide, sulfobromophthalein, and blockers of anion transporters. The anion flux generated by ABC1 is up-regulated by orthovanadate, cAMP, protein kinase A, and okadaic acid. In other ABC transporters, mutating the conserved lysine in the nucleotide binding folds was found to severely reduce or abolish hydrolysis of ATP, which in turn altered the activity of the transporter. In ABC1, replacement of the conserved lysine 1892 in the Walker A motif of the second nucleotide binding fold increased the basal ionic flux, did not alter the pharmacological inhibitory profile, but abolished the response to orthovanadate and cAMP agonists. Therefore, we conclude that ABC1 is a cAMP-dependent and sulfonylurea-sensitive anion transporter.

1-Methyl-3-isobutylxanthine↗

PubMed1997

Low-threshold Na+ currents: a new family of receptor-operated inward currents in mammalian nerve cells.

In the mammalian nervous system, various neurotransmitters can modulate cell excitability by inducing slow membrane potential changes. In the last decade, inhibition of potassium currents has been characterized as the primary mechanism by which neurones can undergo sustained depolarization. More recently (1990s), a new class of inward currents, which are voltage-dependent and mainly carried by sodium ions, has been found to be activated by various neurotransmitter receptors in mammalian central and peripheral neurones. Because the channels involved pass depolarizing current, are open at more negative membrane potentials than the resting potential, and are voltage-gated and persistent, these currents are capable of producing regenerative and maintained depolarizations and play an important role in neuronal signalling.

Action Potentials↗

PubMed1997

[Mammary carcinoma in a patient with hyperprolactinemia].

In 1990, a 50 year-old man was referred to us for hyperprolactinemia. At 37 years of age the patient had undergone left mastectomy, for a histologically confirmed gynecomastia and, in 1989, he had undergone pituitary adenomectomy, for a PRL secreting macroadenoma (PRL = 3520 ng/ml). Persistently high PRL plasma levels (PRL = 550 ng/ml) showed an incomplete surgical removal of the adenoma and as a consequence, radiotherapy of the pituitary area was performed in 1990. When the patient referred to us, PRL plasma levels were still pathologic and medical therapy with bromocriptine was started. A year later a replacement therapy with cortisone, testosterone, L-thyroxine, was commenced, as the patient presented a post-radiotherapy hypopituitarism. Since the treatment with bromocriptine was unsuccessful, the drug was replaced with cabergoline, but not even the latter was able to normalize PRL plasma levels. In 1996, a nodule of 3 cm in diameter was discovered under his right mammary areola. The nodule biopsy showed a grade II infiltrating ductal breast carcinoma positive to the estrogen and progesterone receptors analysis. A right total mastectomy was performed and the diagnosis was confirmed through histological examination. A case of gynecomastia and breast cancer in a male patient who had been exposed to high PRL plasma levels for several years is reported. In this patient, both elevated PRL plasma levels and a relative hyperestrogenic state may have contributed to originate breast cancer.

Adult↗

PubMed1996

Glial potassium channels activated by neuronal firing or intracellular cyclic AMP in Helix.

1. Cell-attached and whole cell patch clamp experiments were performed on satellite glial cells adhering to the cell body of neurones in situ within the nervous system of the snail Helix pomatia. The underlying neurone was under current or voltage-clamp control. 2. Neuronal firing induced a delayed (20-30 s) persistent (3-4 min) increase in the opening probability of glial K+ channels. The channels were also activated by perfusing the ganglion with a depolarizing high-K+ saline, except when the underlying neurone was prevented from depolarizing under voltage-clamp conditions. 3. Two K(+)-selective channels were detected in the glial membrane. The channel responding to neuronal firing was present in 95% of the patches (n = 393). It had a unitary conductance of 56 pS, a Na+ :K+ permeability ratio < 0.02 and displayed slight inward rectification in symmetrical [K+] conditions. It was sensitive to TEA, Ba2+ and Cs+. The following results refer to this channel as studied in the cell-attached configuration. 4. The glial K+ channel was activated by bath application of the membrane-permeant cyclic AMP derivatives 8-bromo-cAMP and dibutyryl-cAMP, the adenylyl cyclase activator forskolin and the diesterase inhibitors IBMX, theophylline and caffeine. It was insensitive to cyclic GMP activators and to conditions that might alter the intracellular [Ca2+] (ionomycin, low-Ca2+ saline and Ca2+ channel blockers). 5. The forskolin-induced changes in channel behaviour (open and closed time distributions, burst duration, short and long gaps within bursts) could be accounted for by a four-state model (3 closed states, 1 open state) by simply changing one of the six rate parameters. 6. The present results suggest that the signal sent by an active neurone to satellite glial cells is confined to the glial cells round that neurone. The effect of this signal on the class of glial K+ channels studied can be mimicked by an increase in glial cAMP concentration. The subsequent delayed opening of the glial K+ channels does not appear to play a role in siphoning the excess K+ released by active neurones. It is hypothesized that the cAMP-gated glial K+ channels may be involved in the control of glial cell proliferation.

Action Potentials↗

PubMed1996

Appearance of neurofilament subunit epitopes correlates with electrophysiological maturation in cortical embryonic neurons cocultured with mature astrocytes.

E14 rat cortical neurons which have almost no glial progenitors were cocultured with a homogeneous population of mature type 1 astrocytes at a 4/1 ratio in serum free medium. Maturation of neurons was evaluated using a set of well characterized antibodies and two new monoclonal antibodies (MN2E4 and MN3H6) raised against various neurofilament subunits and whole-cell patch clamp experiments. We observed that this coculture method leads to a well-timed and very homogeneous neuronal maturation and that sequential appearance of neurofilament subunits in developing neurons correlates with the electrophysiological maturation. This sequence, early expression of the 68 kDa neurofilament subunit and late appearance of the 200 kDa neurofilament subunit, occurs in normal brain development, which validates this culture model as a useful tool for studying neuronal maturation and differentiation. MN2E4 staining (non-phosphorylated 200 kDa cytoskeletal protein antibody) appeared just before the neurons became excitable. It could thus be used as a functional neuronal marker. MN3H6 staining (phosphorylated 160-200 kDa neurofilament subunit antibody) appeared just after the neurons made synaptic contacts and generated synaptically driven spike bursts. This finding indicated that some phosphorylated epitopes of 160-200 kDa neurofilament followed synaptogenesis. These processes may play a key role in stabilizing the synapses to achieve a functional neuronal network.

Animals↗

PubMed1996

Muscarinic activation of a novel voltage-sensitive inward current in rabbit prevertebral sympathetic neurons.

The muscarinic activation of rabbit prevertebral sympathetic neurons was studied in non-dissociated coeliac and superior mesenteric ganglia using whole-cell patch-clamp techniques. In the presence of nicotinic blockers, carbachol, muscarine and oxotremorine-M (1-50 microM) induced tonic firing by activating a persistent inward current. These effects were abolished by atropine. They persisted when the M-current was blocked with Ba2+ (1 mM) and intracellular Cs+. The muscarinic inward current was found to be time- and voltage-dependent. It peaked at -60 mV, decreased at large hyperpolarizations and was tonically activated between -110 and -20 mV, which gave steady-state I-V curves an N-shape between -96 and -54 mV. The negative slope accounted for the large hyperpolarizing responses generated by current pulses in carbachol-treated cells. The muscarinic current was abolished when Na+ was replaced by choline, Tris+, sucrose, N-methyl-D-glucamine and Cs+ but not Li+. It was resistant to tetrodotoxin (3 microM), amiloride (3 microM), benzamil (10 microM) and tetraethylammonium (5-20 mM). No involvement of K+ and Cl- could be detected. We therefore styled it INa,M, in reference to its ionic selectivity and its coupling to muscarinic receptors. Low Ca(2+)-Mg2+ salines enhanced the Na,M-current. The current was blocked by Cd2+, Co2+, La3+ (1 mM) and Ba2+ (5 mM) but insensitive to methoxyverapamil hydrochloride, nicardipine, nifedipine and omega-conotoxin MVII A (2-20 microM). These effects were ascribed to the binding of di- and trivalent ions to the Na,M-channels. Spike bursts transiently blocked INa,M. With high intracellular ethylene glycol bis(b-aminoethyl ether)-N,N'-tetraacetic acid or 1,2-bis (2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (20-50 mM), this effect was reduced, whereas INa,M persisted in long-term recordings and its amplitude increased twofold, indicating that intracellular calcium negatively regulated the Na,M-channels. We conclude that we have described a novel muscarinic receptor-coupled channel which appears to play a major part in regulating the firing behaviour of sympathetic neurons.

Animals↗

PubMed1996

Synaptically activated low-threshold muscarinic inward current sustains tonic firing in rabbit prevertebral sympathetic neurons.

Whole-cell patch-clamp experiments were performed on non-dissociated rabbit coeliac sympathetic neurons in the presence of nicotinic blockers. Coeliac neurons were classified as either silent or spontaneously active (pacemaker) cells. Under voltage-clamp conditions, pacemaker cells exhibited a steady-state N-shaped current-voltage relationship due to the presence of a persistent voltage-dependent inward current in the potential range of -100 to approximately -20 mV. This inward current sustained the regular firing activity of pacemaker cells and was absent from quiescent neurons. It disappeared in the presence of tetrodotoxin and in low Ca(2+)-high Mg2+ external solutions and was enhanced by eserine. Splanchnic nerve stimulation induced slow regenerative depolarizations and firing discharges in silent neurons by activating a low-threshold voltage-sensitive inward current. The synaptic current had a U-shaped voltage-dependence from -96 to approximately -20 mV and exhibited the dynamic properties of the muscarinic voltage-dependent inward current INa,M. It gave the current-voltage relationship an N shape similar to that observed in spontaneously active cells. The muscarinic antagonists atropine and pirenzepine abolished the inward current present in pacemaker cells and that induced by nerve stimulation in silent neurons. These data provide evidence that both spontaneous firing activity and nerve-evoked depolarizing responses in coeliac neurons are sustained by the activation of the muscarinic Na,M current. The tonic activation of INa,M in spontaneously firing cells results from a sustained Ca(2+)-dependent tetrodotoxin-sensitive release of acetylcholine. This study provides evidence that the role of the muscarinic receptors is not purely a neuromodulatory one, but that these receptors are directly involved in ganglionic neurotransmission.

Animals↗

PubMed1996

Influence of urinary calcium concentration on erythrocyte morphology.

The effect of the urinary calcium concentration (CaU) on erythrocyte morphology was studied by incubating erythrocytes in urine with prefixed CaUs of 5, 10, 20 and 40 mmol/l by addition of CaCl2. The same experiment was carried out on erythrocytes preincubated with levo-verapamil (l-V) at 10, 100 and 200 mumol/l. Phase contrast microscopy observations were performed at 0, 30, 60, 120, and 240 min by 2 experienced investigators. At 0 min the erythrocytes showed a clear extraglomerular pattern. At 60 min marked morphological and volumetric alterations were evident when the CaU was > or = 10 mmol/l. On the contrary, no change was found when red cells were treated with > or = 100 mumol/l l-V, independent of the CaU. Dysmorphic erythrocyturia has been related to transglomerular passage even if it was sporadically observed in hypercalciuric or lithiasic patients. This work suggests a role for a high CaU in causing the formation of microcytic and warped erythrocytes. In our opinion, in hypercalciuric urine the appearance of dysmorphic or mixed hematuria does not necessarily indicate transglomerular passage.

Calcium↗

PubMed1996

Antibody-coated electrodes for detecting somatic exocytosis of somatostatin-like material in Helix neurones.

Immunoelectrodes have been developed which can be used to detect minute amounts of somatostatin. They were made with electrochemically treated glassy carbon fibres coated with anti-somatostatin antibodies. Calibration and various controls were carried out to ensure that the immunoelectrodes responded specifically to the presence of femtomolar somatostatin. Electrophysiological experiments were performed on anti-somatostatin immunoreactive neurones in the snails Helix aspersa and H. pomatia. Somatostatin-like material was released in response to sustained firing. The release was measured at the soma, which means that it occurred in the extrasynaptic area. The finding that the fluorescent dye FM 1-43 was incorporated into somatic vesicles confirmed that exocytosis actually occurred during sustained neuronal firing.

Animals↗

PubMed1995

Choline blocks large conductance KCa channels in mammalian sympathetic neurones.

Using the outside-out configuration of the patch clamp technique, we examined the effects of externally applied choline chloride (ChCl) on the large-conductance calcium-dependent K+ (KCa) channel in sympathetic neurones of the rabbit coeliac ganglion. Isotonically replacing the bath sodium chloride by ChCl significantly decreased the unitary current amplitude of the KCa channels but did not affect their gating properties. The blocking effect was dose-dependent and required 24 mM ChCl to reach half-reduction of the KCa channel conductance (from 134 to 64 pS). In whole-cell voltage-clamped neurones, ChCl activated both nicotinic and muscarinic receptors, which induced various membrane current changes including Ca2+ current decrease. We therefore tested the effects of the cholinergic agonists acetylcholine and carbachol on excised KCa channels. Both agents failed to mimic the blocking effects of ChCl. We therefore conclude that KCa channels in sympathetic neurones are occluded by external choline chloride.

Animals↗

PubMed1995

Emergence of a synaptic neuronal network within primary striatal cultures seeded in serum-free medium.

In order to investigate the basic cellular mechanisms involved in neuronal interactions within the striatum, we prepared a primary striatal cell culture from rat fetal brain in chemically defined medium. Using morphological and whole-cell recording methods, we observed that an intensive neuritic elongation with a progressive build up of a sodium-dependent electrogenesis occurred during the first week of culture. Morphologically mature synapses began to develop after 10 days in vitro. By this time, most of the neurons (82 +/- 9%) received spontaneously synaptic potentials, which led them to fire (71 +/- 11%). The spontaneous firing was prevented by cadmium (200 microM) and tetrodotoxin (5 microM), which suggested that a Ca(2+)-dependent release of neurotransmitters was involved in the synaptic activation. We further obtained evidence that GABA, and to a lesser extent acetylcholine, contributed to these spontaneous synaptic potentials. At 15 days in vitro, it was possible to observe up to four synaptic contacts on a given dendrite. By this time, whole-cell recordings performed on pairs of neurons showed that the mature neurons were interconnected by excitatory synapses. As the number of synapses increased, the striatal neurons gradually formed a large network in which spontaneous activity developed, which tended to be organized into synchronized bursting patterns.

Action Potentials↗