Edward S. Redgate

Selective enhancement of the lordotic component of female sexual behavior in rats following destruction of central catecholamine-containing systems

Ovariectomized female rats received two intraventricular injections of 200 microgram of 6-hydroxydopamine (6-OHDA), a treatment which produced 66% depletion of telencephalic norepinephrine and 53% depletion of telencephalic dopamine. Compared to vehicle-injected controls, 6-OHDA-treated animals showed increased lordosis scores when treated with ovarian hormones. This effect was potentiated by additional treatment with 100 mg/kg alpha-methyl-p-tyrosine (AMT), a catecholamine synthesis inhibitor. Besides showing increased frequency and intensity of lordosis, animals treated with both 6-OHDA and AMT retained the lordotic posture significantly longer after the male dismounted than animals given either treatment alone or vehicle controls. The enhancement of lordosis following CA depletion was not prevented by a series of dexamethasone treatments which caused a marked suppression in adrenal steroid (corticosterone) levels. This suggests that normal adrenal function is not a prerequisite for the observed enhancements. It was concluded that the lordotic response is inhibited by the activity of a catecholamine system. Soliciting behavior (hop-darting) was not enhanced by any treatment, suggesting that catecholamine activity has an inhibitory influence on the stop component of sexual behavior, but not on the whole copulatory pattern.

Plasma corticosterone levels during sexual behavior in male rats

The activity of the pituitary-adrenal (p-a) system, as reflected in plasma levels of corticosterone, was determined in 14 male Long Evans rats during copulation, exposure to an open field and in control conditions. Plasma corticosterone concentration during copulation was higher than in control conditions (13.3 ± 1.2 vs 9.4 ± 1.2 μm%, p < .01), but well below mean levels obtained in the open field (21.2 ± 0.8 μm%). Individual data indicated that some males gave no evidence of p-a activation during sexual activity. Furthermore, animals which showed increased steroid levels during copulation tended to have longer latencies to reinitiate copulation after ejaculation and were behaviorally less active in a subsequent open field test. It was suggested that neither sexual arousal nor copulatory performance necessarily activates the p-a system. Males showing p-a activation may be slow to habituate to a novel stimulus and thus the elevated steroid levels may reflect an insufficient number of habituation trials with the receptive female.

Polyamines in brain tumor therapy

SummaryIn the search for ways to augment current brain tumor therapies many have sought to exploit the fact that adult brain tissue is virtually lacking in cell division. This endorses a special appeal to therapeutic approaches which target the dependence on cell division for brain tumor growth. Polyamines play an essential role in the proliferation of mammalian cells and depletion results in inhibition of growth. As a result, there are investigations into the feasibility of controlling tumor growth by targeting the enzymes in polyamine metabolism with specific enzyme inhibitors. DFMO, an inhibitor of putrescine synthesis, is a cytostatic agent which in combination with tritiated radioemitters or cytotoxic agents such as, MGBG or BCNU is an effective antitumor agent, but the effectiveness of DFMOin vivo is reduced by tumor cell uptake of polyamines released into the circulation by normal cells and from gut flora or dietary sources. However, DFMO therapy combined with elimination of exogenous polyamines inhibits tumor growth but also results in body weight loss, reduced protein synthesis and evidence of toxicity. Furthermore, tumor growth recurs upon termination of treatment. In contrast, competitive polyamine analogs function in the homeostatic regulation of polyamine synthesis but fail to fulfill the requirements for growth and they continue to inhibit tumor growth for several weeks after cessation of treatment. Analogs are now in clinical trials. However, their action may be highly specific and differ from one cell type to another. We suggest that the effectiveness of polyamine based therapy would be enhanced by two approaches: local delivery by intracerebral microdialysis and tumor cell killing by internal radioemitters such as tritiated putrescine or tritriated thymidine which are taken up in increased amounts by polyamine depleted tumor cells. The growth inhibition by polyamine depletion prevents the dilution of the radioactive putrescine and thymidine. The overload of radioactivity kills the growth inhibited cells so that growth cannot recur when treatment terminates.

Effects of immobilization stress on open field behavior and plasma corticosterone levels of aging C57BL/6J mice

Four age groups of C57BL/6J mice (2.2, 6.2, 12.0, and 23.3 months) were subjected to either immobilization or handling (control) procedures. Open field behavior was observed before and after experimental treatments and plasma corticosterone levels were assessed 11 days following the immobilization or handling procedures. Eleven days following immobilization elevated corticosterone levels were observed for all but the 12.0 month group of mice. No behavioral effects were observed for the experimental groups, although both locomotor activity and exploratory behavior declined with advancing age. The age-related decrease in activity was entirely accounted for by scores on the initial open field test. Exploratory behavior was observed to be a more complex function of both age and experience.

A new rat brain tumor model: Glioma disseminated via the cerebral spinal fluid pathways

SummaryA rat brain tumor model has been developed with the clinical and pathological features of dissemination via the cerebral spinal fluid (CSF) pathways. A precise number of 9L gliosarcoma cells (5 × 102 to 5 × 105) is stereotactically injected into the CSF of the lateral ventricle. The interval until the onset of neurological symptoms and then death is reproducible and dependent upon the number of cells injected. The median survival of three groups of rats receiving 5 × 105 cells in three different experiments was 17, 18 and 19 days respectively. For three groups receiving 5 × 104 cells, the median survival was 23, 24 and 25.5 days respectively and for two groups receiving 5 × 103 cells the median survival was 28 and 30 days respectively. The animals developed multiple tumor implants along the CSF pathways usually resulting in hydrocephalus. This tumor model was developed to simulate dissemination via CSF pathways as seen with medulloblastoma and other primitive neuroectodermal tumors of the central nervous system. It will be used to evaluate the therapeutic efficacy of intra-ventricularly administered anti-neoplastic drugs against small implants and malignant cells in the CSF pathways.

Interrelationship of plasma cortisol and other activation indices during EMG biofeedback training

Plasma cortisol, cephalic electromyography (EMG), heart rate, fingertip vasoconstriction/dilation, respiration rate, self-reported anxiety, and target symptom severity were monitored in 24 human outpatients who volunteered to undergo the novel experience of an eight-session EMG biofeedback-based relaxation training program. Acute cortisol levels were generally found to be positively related to heart rate, degree of vasoconstriction, and self-reported anxiety but independent of cephalic EMG level and respiration rate. For the 12 participants above the median in initial trait anxiety, the mean reductions in cortisol were 22, 19, and 31% (relative to baseline) at the fourth, eighth, and 1-month follow-up sessions, respectively. Although there was some indication of the presence of a centrally integrated state of lowered neurophysiological arousal, the multiple dependent measures suggest that “relaxation” in this situation is not necessarily a simple unitary physiological and psychological event.

Intra-cerebral ventricular infusion of 5-IODO-2-deoxyuridine (IUDR) as a radiosensitizer in the treatment of a rat glioma☆

The efficacy of 5-iodo-2-deoxyuridine (IUDR) as a radiosensitizer when administered by continuous infusion into the cerebral spinal fluid (CSF) of the lateral cerebral ventricle was evaluated in a 9L gliosarcoma rat brain tumor model. Stereotactic implantation of a 5 x 10(4) tumor cell suspension into the left caudate nucleus was carried out in four groups of 10 rats each. Control animals had a median survival of 16.9 days (range 16-21 days). IUDR, 8.4 mg over 7 days administered by continuous infusion into the left lateral ventricle produced a slight survival advantage (median survival 21.5 days, range 12-56). Irradiation of the entire brain, 8 Gy on days 4, 6 and 7 after tumor cell implantation also produced a slight improvement in survival (median 19.5 days, range 17-34). The combination of radiation and IUDR infusion into the CSF produced a marked survival advantage (median 30.5, range 22-54) compared to the control and single modality treatment groups. This is the first demonstration of the effectiveness of IUDR as a radiosensitizer when administered into the lateral cerebral ventricle in the treatment of an intraparenchymal brain tumor.

Central nervous system mediation of pituitary adrenal rhythmicity

Abstract Evidence favoring the view that there is a dissociation between the two aspects of pituitary adrenal operation, the stress and the rhythm modes, is as follows 1) it is possible to abolish rhythmic function by anterior hypothalamic lesions without altering the responses to stress; 2) during maturation of the rat CNS, the stress response appears prior to weaning while pituitary-adrenal rhythm appears after weaning; 3) appropriate neuropharmacologic treatment can block circadian rhythm without altering the stress response; 4) responsiveness to ether or immobilization may be independent of the diurnal rhythm; 5) the stress response to a very strong stimulus, such as immobilization, habituates while the circadian rhythm does not. On the other hand there is some evidence in favor of interdependence between the two modes of operation: 1) input channels for stress and rhythm may overlap since strong synchronizers of rhythm, such as food and water restriction and light can become stress stimuli; 2) steroid feedback can alter the timing of the circadian rhythm; 3) circadian variation has been observed in the responses to certain stimuli; 4) neurotransmitters which are prominently implicated in the circadian rhythm also appear involved in the stress response.

Difluoromethylornithine enhanced uptake of tritiated putrescine in 9L rat brain tumors

Difluoromethylornithine (DFMO) depletes endogenous putrescine and enhances the uptake of and retention of [3H] putrescine in vitro. To determine if DFMO also enhances uptake of [3H] putrescine in vivo, DFMO and trace doses of [3H] putrescine, dissolved in artificial CSF, were infused into growing (6-9 day) 9L brain tumors by means of osmotic pumps. When 7-day osmotic pumps were loaded with 1 microCi [3H] putrescine, with or without 10 or 100 mM DFMO, pumped at 1 microl/h, the mean uptake after 3 days was 168 +/- 62 cpm/mg tumor (17 rats) without DFMO, 300 +/- 197 cpm/mg tumor (11 rats) with 10 mM DFMO and 1088 +/- 421 cpm/mg tumor (11 rats) with 100 mM DFMO (p < or = 0.05 vs. control). Significantly less radioactivity was detected in the contralateral brain and in nonbrain tissues (0.5 +/- 0.1 to 14 +/- 5 cpm/mg). To measure the extent of [3H] putrescine distribution in the tumor, the same dose of drugs was delivered for a longer period of time, using 14-day pumps to allow tumors to become large enough to be divided into 1.4 mm thick transections. The mean radioactivity in the sections from eight control rats receiving [3H] putrescine without DFMO were not significantly different between the sections (174 +/- 61 cpm/mg tumor for sections containing the cannulas, 273 +/- 61 and 259 +/- 91 cpm/mg for adjacent sections). In the six rats given 100 mM DFMO there was a significant increase in mean radioactivity in the cannula containing section (2251 +/- 919 cpm/mg tumor). Mean counts from adjacent sections in these rats were 97 +/- 44 and 33 +/- 13 cpm/mg. Values for contralateral corpus striatum and nonbrain tissues ranged from 0.7 +/- 0.3 to 4.3 +/- 1.5 cpm/mg tissue. When DFMO was delivered directly to the tumors while [3H] putrescine was infused intraperitoneally, the uptake in the tumor slices was low (5-10 cpm/mg in different slices). These results demonstrate that infusion of DFMO directly into growing 9L brain tumors can selectively enhance the uptake of exogenous [3H] putrescine by rapidly dividing cells which are within a 1.4 mm diameter area at the cannula tip. Although these studies used [3H] putrescine at trace doses, it is estimated that infusion of higher doses of [3H] putrescine plus DFMO will selectively kill tumor cells.

Effect of D,L-α-difluoromethylornithine (DFMO) enhanced [3H] putrescine uptake on 9L tumor cell growth and colony forming efficiency☆

PURPOSE This study explored the possible use of D,L- alpha-difluoromethylornithine (DFMO) to enhance the uptake of [3H] putrescine in order to selectively kill brain tumor cells. METHODS AND MATERIALS Gliosarcoma cells (9L) were grown for 4 or 20 day periods in monolayer cultures with or without [3H] putrescine and/or DFMO. Cells in culture incubated for 20 days were replated at 4-day intervals. Cells were counted on a Coulter Electronic Particle Counter and percent viability was determined by eosin dye exclusion. Survival of cells with proliferative capacity was assayed by their colony. Forming ability and surviving fraction was calculated. The radioactive counts due to [3H] putrescine were measured in 9L cells and in medium and expressed as cpm/100 cells or cpm/ml, respectively. RESULTS As previously reported (15), DFMO treatment resulted in termination of cell proliferation that was reversible by the addition of exogenous putrescine. Specifically, after 4 days in culture, cell counts in groups exposed to 10 mM DFMO were 55% of those in control groups and addition of 3 mM putrescine reversed the DFMO effects. Uptake of [3H] putrescine into untreated cells increased in proportion to the amount of exogenous putrescine present during 4 days of culture (range 0.01 nmol to 100 nmol) and the presence of DFMO in the medium enhanced the uptake 9 fold throughout these ranges. At activities greater than 100 cpm/100 cells the cell count was reduced to 23 to 48% of control after 4 days in culture. Extending the treatment to 20 days of incubation increased the killing of 9L cells. During the 20-day incubation, control cells increased from 5 x 10(5) to 13 x 10(12) of which 90% were colony forming cells. Treatment with either 25 microCi [3H] putrescine or 1 mM DFMO for 4 days followed by removal of these agents and incubation for an additional 16 days for a total of 20 days resulted in 31 x 10(8) or 18 x 10(7) colony forming cells, respectively. Combining [3H] putrescine and DFMO treatments during the first 4 days of the 20 day incubation reduced the colony forming cells to 21 x 10(5) (surviving fraction to 67%). When the DFMO treatment was present during the entire 20 days, it became cytotoxic since the colony forming cells were reduced to 35 x 10(3) (surviving fraction was 17%). The combination of the 4-day [3H putrescine and the 20 day DFMO treatments resulted in only 1200 surviving colony forming cells (surviving fraction was only 2%). CONCLUSION DFMO treatment of 9L cells for 20 days resulted in increased uptake of [3H] putrescine, a 10(10) fold inhibition of colony forming cells and extensive 9L cell killing relative to untreated controls.