Brevetoxin-induced neural insult in the retrosplenial cortex of mouse brain

Brevetoxins (polyether breve toxins; PbTx) are polyether neurotoxins produced by the marine dinoflagellate Karenia brevis, an organism associated with red tide blooms in the Gulf of Mexico and along the Atlantic coast from Florida to North Carolina. Brevetoxin-3 (PbTx-3) is a major component of the array of brevetoxins found in marine aerosols measured along red tide affected beaches. Humans exposed to aerosolized brevetoxins for short periods of time often suffer a variety of adverse health effects. It was consequently of interest to assess the potential for aerosolized brevetoxin to produce a neurotoxic response. Female BALB/c mice were exposed nose-only for 2 consecutive days to PbTx-3 aerosol, with a 2-h exposure on the first day and a 4-h exposure on the second day. The average PbTx-3 exposure concentrations on days 1 and 2 were 312 +/- 113 mug brevetoxin 3/m3 and 278 +/- 24 mug brevetoxin 3/m3, respectively. The brevetoxin-containing aerosol had a mass median aerodynamic diameter of 0.92 mum with a geometric standard deviation of 1.38. Coronal sections of mouse brains were evaluated for neuronal damage using both silver and Fluoro-Jade B staining to identify degenerating neuronal elements. PbTx-3 inhalation exposure produced neuronal degeneration in the posterior cingulate/retrosplenial cortex of mice as evidenced by silver-positive degenerating neurons in this region. No staining was found in other regions of the PBTx-3-exposed mouse brains or in brains of control, sham-exposed mice. The existence of a neurotoxic insult in PbTx-3-exposed mice was confirmed using Fluoro-Jade B to label degenerating neurons. Fluro-Jade-positive neurons were observed in the retrosplenial cortex of PBTx-3 exposed, but not control, mice. These results suggest that subacute exposure to PbTx-3 for 2 days is sufficient to induce neuronal degeneration in a discrete region of the mouse cerebral cortex.     

Effect of desipramine on inositol phosphate formation and inositol phospholipids in rat brain and human platelets.

To examine the mechanism of action of antidepressant drugs, we studied the effect of desipramine (DMI) in vitro on agonist-stimulated inositol phosphate formation and inositol phospholipids in rat brain and human platelets. We observed that DMI inhibited thrombin-stimulated 3H-inositol bisphosphate (IP2) and 3H-inositol trisphosphate (IP3) but not 3H-inositol monophosphate (IP1) formation in human platelets. DMI also inhibited norepinephrine (NE) and serotonin (5-HT) stimulated 3H-IP1 formation in rat cerebral cortex. DMI increased levels of all three 3H-inositol phospholipids, 3H-phosphatidyl inositol (PI), 3H-PI-4-phosphate (PIP), and 3H-PI 4,5-bisphosphate (PIP2), in both platelets and rat cortex. The decreased formation of inositol phosphates and increased levels of [3H]-PI, [3H]-PIP, and [3H]-PIP2 by DMI appears to be due to the inhibition of the enzyme phospholipase C rather than its effects on receptors. It is thus possible that interaction of tricyclic antidepressant drugs with the PI-signaling system may be related to their mechanism of action.     

Diphenylhydantoin binding to brain lipids and phospholipids.

Partition coefficients for diphenylhydantoin (DPH) were determined using four organic solvents and a 0.1 M phosphate buffer. Values ranged from 25.5 for chloroform to 0.02 for hexane. When chloroform-methanol-soluble brain lipids were added, there was a marked enhancement of DPH entry into hexane; 2 mg lipid resulted in an almost equal distribution between hexane and buffer. Proteolipids produced a similar but quantitatively small change. Neutral fat, cholesterol and glycolipids w had no effect on DPH distribution. A number of commercially available phospholipids were tested and all increased the hexane/aqueous partition coefficient of DPH, although there was considerable variation among the several phospholipids employed. Neither total DPH concentration nor the addition of cations influenced this distribution. These results provide strong evidence for binding of DPH to phospholipids.     

Effect of choline on tetraethylammonium transport in mouse kidney cortex slices

Choline inhibits the uptake of [¹⁴C]tetraethylammonium (TEA) by mouse kidney cortex slices incubated in Krebs-Ringer bicarbonate buffer (37°, pH 7.4)), aerated with O₂-CO₂ 95:5 v/v%. Inhibition seemed to be of a competitive type. Stimulation of unidirectional [¹⁴C]TEA exit occurred when choline was added to the wash-out medium, and approached 'a maximum value with increasing choline concentrations. Enhancement of TEA exit by choline could be demonstrated in the presence of high external K⁺ concentrations suggesting that it was not secondary to unspecific changes in the membrane potential. Initial [¹⁴C]TEA uptake was increased in slices preloaded with choline. This argues against the possibility that choline stimulates exit by displacing [¹⁴C]TEA from intracellular binding sites. Preloading the tissue with choline did not reduce relative [¹⁴C]TEA exit which indicates that stimulation of initial TEA uptake after choline preloading is due to an increase of TEA influx rather than to an inhibition of TEA efflux. The results suggest that choline and TEA share a common transport mechanism. Transfer of TEA across the membrane in both directions depends on the transconcentration of choline. The stimulation phenomenon may be an example of accelerative exchange diffusion.     

Effect of escalating doses of d-fenfluramine on the content of indoles in brain.

The neurochemical effects of a large dose challenge (5 mg/kg, i.p.) of d-fenfluramine (d-F) in rats, given saline or gradually escalating doses of d-F (0.1-2.5 mg/kg, i.p.), were examined with regard to regional sensitivity and the time-course of recovery. The indole-depleting effect after the large dose of d-F to saline-pretreated animals appeared to differ, depending on the areas of brain considered (cortex greater than hippocampus greater than striatum), despite the fact that the drug and its main metabolite, d-norfenfluramine (d-NF) distributed almost uniformly in the regions of brain examined. The depletion in all these regions of the brain was reversible within 6 weeks, serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) being back to control levels in the hippocampus and striatum but not 5-HT in the cortex. However, when rats were exposed to gradually escalating doses of d-F the recovery of indoles in the brain, after injection of the large dose challenge, appeared to be faster. Indoles were markedly less reduced 1 week later in the cortex, hippocampus and striatum, with content of indole in the striatum showing complete recovery and the long-term depletion of 5-HT and 5-HIAA, by the subsequent large dose challenge was almost completely reversed in all regions. Analysis of the concentrations of d-F and its main metabolite d-fenfluramine (d-NF) in brain excluded any pharmacokinetic tolerance. These results suggest that during therapeutic treatment with d-F, the use of escalating doses may attenuate the potential for the long-lasting decrease of 5-HT in brain.     

Effect of procarbazine on benzodiazepine receptor in mouse brain

Procarbazine, a non-benzodiazepine tranquilizer and anti-convulsant, significantly decreased the binding of [3H]diazepam to mouse cerebellar membranes. The drug treatment reduced both the affinity and density of the receptor. The effect, however, was more pronounced on the number of binding sites suggesting occupation of the receptor by procarbazine. The results suggest that benzodiazepine receptors may also be involved in the pharmacological action of non-benzodiazepine drugs.     

Effect of anti-thyroid agents, methimazole and propylthiouracil, on brain noradrenaline content.

1 Methimazole (1-methyl-2-mercaptoimidazole, MMI) and propylthiouracil (6-propyl-2-thiouracil, PTU) which are used in the therapy of hyperthyroidism were found to reduce brain noradrenaline (NA) content. Endogenous NA levels in rat brain were reduced from 1 to 6 h after intraperitoneal injection of MMI by doses in excess of 25 mg/kg and by PTU at a dose of 50 mg/kg. However, endogenous NA in the rat heart was only slightly reduced after 50 mg/kg of MMI, and was not affected by PTU (50 mg/kg). 2 Both MMI and PTU effectively inhibited the in vivo conversion of [3H]-dopamine into [3H]-noradrenaline ([3H]-NA) in the brain of rats after a single intraperitoneal injection of doses above 10 mg/kg (MMI) and 25 mg/kg (PTU). This inhibition by MMI and PTU was dose-dependent over the range of 10 mg/kg to 50 mg/kg, was highest after 2-3 h and continued for at least 6 h after their injection; The conversion rates returned to normal after 24 hours. 3 The results suggest that the reduction of brain NA by these drugs is, at least in part, due to the inhibition of brain dopamine beta-hydroxylase.     

褪黑激素对小鼠脑细胞膜流动性及丙二醛含量的影响

以荧光染料DPH负载脑皮层突触体或细胞, 用MPF-4荧光分光光度计检测膜流动性; TBA比色法测定丙二醛(MDA)含量,观察褪黑激素(MT)对小鼠脑细胞膜流动性和MDA含量的影响. 结果显示MT(1或20 mg·L^-1饮水中口服3个月,最后1个月再ip MT 0.5或2.0 mg·kg^-1·d^-1)能够提高老年小鼠脑细胞膜的流动性,MT 0.01-1.0 μmol·L^-1拮抗氧化剂诱发的分离新生小鼠脑细胞膜流动性下降及部分拮抗其MDA含量增加. 结果说明,作为一种体内抗氧化激素, MT提高脑细胞膜流动性,降低脑细胞内MDA含量, 从而对神经元有保护作用,是其重要的抗衰老机理之一.