Donna Schorer-Apelbaum

Effect of prenatal stress on plasma corticosterone and catecholamines in response to footshock in rats

The effect of prenatal stress was investigated on the sympathoadrenal response to novelty and footshock by measuring the time course of the changes in circulating corticosterone (COR) catecholamines and their metabolites. Pregnant rats were subjected to noise and light stress, three times weekly on an unpredictable basis throughout gestation. When the male offspring of stressed rats (PS) and those of unstressed mothers (C) were 4.5-5 months of age, they were prepared with indwelling catheters in the tail artery 24 h before the experiment. Resting levels of plasma COR, noradrenaline (NA), adrenaline (AD), dihydroxyphenylglycol (DHPG), dihydroxyphenylacetic acid (DOPAC), and dihydroxyphenylalanine (DOPA) were measured. Further blood samples were taken within 3 min of their transfer to the shock box, 1-2, 5, 15, and 45 min after footshock. Plasma COR was significantly higher in PS than in C rats at rest, but those of adrenaline, NA, and their metabolites did not differ in the two groups. Transfer of the rats to the shock box increased plasma COR, NA, adrenaline, and dihydroxyphenylglycol in both groups, and dihydroxyphenylalanine and dihydroxyphenylacetic acid only in PS rats. All the catechols increased further 2-3 min after footshock, except dihydroxyphenylalanine in PS rats. Plasma NA and dihydroxyphenylglycol levels were significantly higher in PS than in C rats immediately after footshock, indicating a greater activation of the sympathetic nervous system in PS rats. The findings demonstrate for the first time that prenatal stress can induce long term changes in the sensitivity of the sympathoadrenal system to stress.

Gender differences in sympathoadrenal activity in rats at rest and in response to footshock stress

A comparison was made of the dynamics of sympathoadrenal activity in 11 age‐matched male and female rats, under basal conditions and after exposure to footshock. Rats were prepared with indwelling catheters in the tail artery 24 h before the experiment. Measurements were made of plasma corticosterone (COR), norepinephrine (NE), epinephrine (EPI), dihydroxyphenylalanine (DOPA), dihydroxyphenylglycol (DHPG) and dihydroxyphenylacetic acid (DOPAC) under resting conditions, after transfer to the shock box (novelty) and at various times after footshock. Under basal conditions, males have significantly higher blood pressure and plasma DHPG/NE ratios but lower plasma levels of COR, NE and DOPAC than females. Three min after exposure to the shock chamber (novelty stress) there were significant increases in COR, EPI, NE and DHPG in both sexes, while DOPA increased only in females and DOPAC remained unchanged in both sexes. Footshock produced a further increase in EPI, NE and DOPAC within 2 min, which lasted about 15 min. There were significant sex differences in the extent and duration of the response of COR, EPI and DHPG. The data show that the female sympathoadrenal system is more reactive than that of the male to the stresses of a novel environment and footshock. The smaller DHPG/NE ratios in females at rest and after stress suggest that neuronal uptake of NE is lower in females than in males. The finding that stress produces larger increments of plasma DOPA and DOPAC in female rats indicates that tyrosine hydroxylase in the sympathetic nerve terminals and adrenal medulla may also be higher than in males.

Ladostigil prevents gliosis, oxidative–nitrative stress and memory deficits induced by intracerebroventricular injection of streptozotocin in rats

Glial activation and oxidative-nitrative stress occur at an early stage in Alzheimers disease (AD). In a rat model of AD, deficits in cerebral glucose utilization and memory were seen 3-4 weeks after intracerebroventricular (icv) injection of streptozotocin (STZ). This study examined whether icv STZ induced glial activation and oxidative-nitrative stress preceded the memory deficits and whether they could be prevented by ladostigil a novel drug, a cholinesterase and monoamine oxidase inhibitor with neuroprotective activity. One week after STZ injection activated microglia and astrocytes were seen in the cortex, around the cannula penetration area, in the hippocampal CA1 region, corpus callosum, medial and lateral septum. The activated astrocytes showed a significant increase in nitrotyrosine immunoreactivity, a measure of oxidative-nitrative stress. Only 3 weeks later were deficits in episodic (object recognition test) and spatial memory (place recognition) seen in STZ-injected rats. Daily oral administrations of ladostigil (1mg/kg) for 1 week, before and after STZ prevented the glial changes, increase in nitrotyrosine immunoreactivity and memory deficits. Taken together the data support the role of glial activation and oxidative-nitrative stress in discrete brain areas in the aetiology of memory deficits and indicate a potential mechanism for their prevention by drug treatment.

Corticosterone mediates some but not other behavioural changes induced by prenatal stress in rats.

The effect of daily varied stress from days 13–21 of gestation in Wistar rats was investigated by tests of learning and memory and anxiogenic behaviour in the 60‐day‐old offspring of both sexes. Prenatal stress decreased the anogenital distance in males at 1 day of age. Anxiogenic behaviour in the elevated plus maze was seen in prenatally‐stressed rats of both genders. There was no significant gender difference in the rate of spatial learning in the Morris water maze but prenatal stress only slowed that of males. In the object recognition test with an inter‐trial interval of 40 min, females but not males, discriminated between a familiar and novel object. Prenatal stress did not affect object discrimination in females but feminised that in males. Maternal adrenalectomy with replacement of basal corticosterone levels in the drinking fluid prevented all of the above effects of prenatal stress in the offspring. To mimic the peak corticosterone levels and time course of elevation in response to stress, corticosterone (3 mg/kg) was injected twice (0 and 30 min) on days 13–16 and once on days 17–20 of gestation to adrenalectomised mothers. This treatment re‐instated anxiogenic behaviour similar to that induced by prenatal stress, indicating that it is mediated by exposure of the foetal brain to raised levels of corticosterone. However, steroid administration to adrenalectomised dams did not decrease anogenital distance, feminise object recognition memory or slow spatial learning in their male offspring. The findings indicate that other adrenal hormones are necessary to induce these effects of prenatal stress.

The kinetics of inhibition of human acetylcholinesterase and butyrylcholinesterase by two series of novel carbamates

Controlled inhibition of brain acetyl- and butyrylcholinesterases (AChE and BChE, respectively) and of monoamine oxidase-B (MAO-B) may slow neurodegeneration in Alzheimers and Parkinsons diseases. It was postulated that certain carbamate esters would inhibit AChE and BChE with the concomitant release in the brain of the OH-derivatives of rasagiline or selegiline that can serve as inhibitors of MAO-B and as antioxidants. We conducted a detailed in vitro kinetic study on two series of novel N-methyl, N-alkyl carbamates and compared them with rivastigmine, a known anti-Alzheimer drug. The rates of carbamylation (ki) and decarbamylation (kr) of recombinant human AChE were mainly determined by the size of the N-alkyl substituent and to a lesser extent by the nature of the leaving group. ki was highest when the alkyl was methyl, hexyl, cyclohexyl, or an aromatic substituent and lowest when it was ethyl. This suggested that ki depends on a delicate balance between the length of the residue and its degree of freedom of rotation. By contrast, presumably because of its wider gorge, inhibition of human BChE was less influenced by the size of the alkyl group and more dependent on the structure of the leaving group. The data show how the degree of enzyme inhibition can be manipulated by structural changes in the N-methyl, N-alkyl carbamates and the corresponding leaving group to achieve therapeutic levels of brain AChE, BChE, and MAO-B inhibition.

Testosterone mediates sex difference in hypothermia and cholinesterase inhibition by rivastigmine

The fall in body temperature and inhibition of hypothalamic cholinesterase induced by rivastigmine (a pseudo-reversible carbamate inhibitor) were compared in male and female rats. In males, 1.5 mg/kg lowered body temperature by 1 degrees C and in females by 3.2 degrees C (P<0.001) and inhibited cholinesterase by 65% and 74%, respectively (P<0.05). Pilocarpine (20 mg/kg) decreased body temperature by 1.1 degrees C in males and 1.9 degrees C in females (P<0.05). Orchidectomy, but not ovariectomy, abolished the sex difference in the hypothermic effect of pilocarpine and the enzyme inhibition induced by rivastigmine, but not in its effect on body temperature. Testosterone (10 mg/rat) decreased the cholinesterase inhibition and the temperature reduction induced by rivastigmine in gonadectomised males and females, but that induced by pilocarpine in males only. In conclusion, rivastigmine causes less inhibition of cholinesterase because testosterone may interfere with its entry into the brain. Testosterone may further decrease the temperature-lowering effect of rivastigmine and acetylcholine receptor agonists in males by an action at a receptor level.

Endogenous opiates mediate cardiac sympathetic inhibition in response to a pressor stimulus in rabbits.

Two different patterns of response to a pressor stimulus occurred in conscious rabbits. This difference was not apparent when a depressor stimulus was applied. At levels of mean arterial pressure exceeding 120 mmHg one group of animals exhibited a marked bradycardia which was due to sympathetic inhibition in addition to vagal activation while this sympathetic component appeared to be lacking in the second group of animals. Naloxone (0.1 mg/kg i.v.) markedly reduced the sympathetic inhibition elicited by phenylephrine but had no significant effect on the reflex vagal stimulation. Naloxone thereby abolished the difference in sensitivity of baroreflex control of heart rate in response to a pressor stimulus between the two groups of rabbits. Naloxone did not influence the sensitivity of the reflex response to nitroprusside. Morphine (2 mg/kg) increased the vagal component of the baroreceptor reflex in response to a pressor stimulus and the sensitivity of the reflex response to nitroprusside in all the rabbits, and this was antagonized by naloxone (0.1 mg/kg). Morphine also potentiated and naloxone antagonized the bradycardic response at levels of MAP exceeding 120 mmHg, in those rabbits which appeared to lack the cardiac sympathetic inhibitory component of the reflex. The results show that endogenous and exogenous opiates can increase the reflex bradycardia in response to a pressor stimulus in the conscious rabbit. The difference in baroreflex sensitivity in different animals may result from their varying ability to activate endogenous opioid systems which depress cardiac sympathetic activity.

Synthesis and in vitro evaluation of anti-inflammatory activity of ester and amine derivatives of indoline in RAW 264.7 and peritoneal macrophages

UNLABELLED A prolonged increase in pro-inflammatory cytokines, TNF-α and IL-6 occurs in inflammatory diseases. Although existing therapies like steroids and TNF-α antagonists are effective they may cause serious adverse effects. We describe the preparation and evaluation for anti-inflammatory activity of 11 novel derivatives of indoline carbamates with a propionic ester, 2-aminoethyl, 3-aminopropyl 2-(dimethylamino)ethyl or 3-(dimethylamino)propyl group in positions 3 or 1. Compounds 25, 26 and 29 were previously shown to inhibit acetylcholinesterase with IC50s ranging from 0.4 to 55μM and to prevent cytotoxicity induced by reactive oxygen species in a concentration range of 100pM-1μM. Compounds 25, 26, 29, 9, 10, 17 and 18, reduced NO, TNF-α and IL-6 at concentrations of 1-10pM in LPS-activated RAW-264.7 and mouse peritoneal macrophages. The reduction in cytokines by compound 25 was associated with an increase in IκBα degradation and a decrease in the phosphorylation of p38 but not that of ERK. CONCLUSION Indoline derivatives substituted at position 3 with chains carrying ester or amino groups may have potential for the treatment of chronic inflammatory and neurodegenerative diseases.

Sympathetic component of baroreflex control of heart rate is impaired in hypertension-prone (SBH) Sabra rats.

Baroreflex control of heart rate in response to phenylephrine was studied in conscious Sabra hypertension-prone (SBH) rats, at a prehypertensive stage, and hypertension-resistant (SBN) rats. Baroreflex sensitivity as determined from the slope of the relationship of mean arterial blood pressure and heart period was significantly lower in SBH rats (0.58 +/- 0.06 versus 1.71 +/- 0.11 ms/mmHg in SBN rats, P less than 0.01) before the development of hypertension. Sympathetic nerve blockade with guanethidine (15 mg/kg) significantly reduced the slope of the mean arterial blood pressure-heart period relationship in SBN rats to 0.45 +/- 0.05 ms/mmHg (P less than 0.01) and increased the pressor response to phenylephrine, without having any effect on these parameters in SBH rats. Atropine methyl nitrate (1 mg/kg) abolished reflex vagal bradycardia in response to phenylephrine in both groups of rats. This suggests that SBH rats are unable to withdraw the sympathetic cardiac component of the baroreflex in response to a pressor stimulus and appear to rely only on increased vagal activity to effect bradycardia.

Novel indoline derivatives prevent inflammation and ulceration in dinitro-benzene sulfonic acid-induced colitis in rats

BACKGROUND In search of safer treatments for inflammatory bowel disease in subjects not responding to, or showing adverse effects to TNF-α antagonists, we tested three novel indoline carbamates in the 2,4-dinitrobenzene sulfonic acid (DNBS) model of colitis in rats. The compounds have anti-inflammatory activity in other disease models in mice. METHODS AN827 (3-(2-(methoxy carbonyl) ethyl) indolin-4-ylethyl methyl) carbamate (0.1 or 1mg/kg), AN680 (3-(2-(methoxy carbonyl) ethyl) indolin-6-ylethyl methyl) carbamate (1.25 or 2.5mg/kg) and AN917 (3-(3-amino propyl) indolin-4-ylethyl methyl) carbamate (1 or 2mg/kg), 5-aminosalycilic acid (5-ASA) (1 or 100mg/kg) or saline (1ml/kg) were administered rectally 1h after intracolonic administration of DNBS, (35mg/kg in 30% alcohol). Disease severity was assessed four days after DNBS administration by change in body weight, colon weight, area of ulceration, myeloid peroxidase (MPO) activity, colonic TNF-α, IL-6 and IL-1β levels. Histopathological scoring was performed after staining colon sections with hematoxylin and eosin and with antibodies to CD68 and CD11b. RESULTS AN827 (0.1 and 1mg/kg), AN680 (2.5mg/kg) and AN917 (2.0mg/kg) significantly reduced all macroscopic and microscopic parameters of colitis, colonic pro-inflammatory cytokines, TNF-α, IL-1β and IL-6 and MPO activity by about 80%. CONCLUSIONS The indoline derivatives largely prevented the symptoms of colitis and were 500-50 times more potent and more effective than 5-ASA. It may be worth evaluating them in models of established colitis. Since AN827 is strongly bound by plasma proteins no adverse effects are expected if compound is absorbed into the circulation after rectal administration.