Time dependent protection of amifostine from renal and hematopoietic cisplatin induced toxicity.

Efficacy of chemotherapy may be maximized and its toxicity can be minimized if drugs would be administered at specified daily times. The present study was aimed to examine if the protection of amifostine against cisplatin toxicity is time dependent. Amifostine is an organic thiophosphate that protects selectively normal tissues, but not tumors, against the cytotoxicity of DNA binding chemotherapeutic agents such as cisplatin. ICR male mice which were entrained to Light:Dark (L:D) 14:10 were injected (intrapritoneal bolus) for 5 consecutive days with either: cisplatin, cisplatin plus amifostine (administered 30 minutes prior to cisplatin). Injections were given at either 08:00, 13:00, 20:00 or 01:00. Five days later, on day 10, each set of mice was sacrificed (at the same hour corresponds to the injection hour), blood count, blood creatinine and blood urea nitrogen (BUN) were assayed. Cisplatin treated mice exhibited nephrotoxicity, as indicated by increased blood urea nitrogen values and by high blood urea nitrogen to creatinine ratios, as well as myelotoxicity that was indicated by low levels of hemoglobin and platelets. Co-administration of amifostine-cisplatin reversed both, the nephrotoxicity of cisplatin, and its myelosuppressive effects. For BUN, hemoglobin and platelets, maximal protections were observed at 08:00, (p <0.05, p <0.01 and p <0.01 respectively). For BUN/Cr ratio (p <0.05), maximal protections was observed at 13:00. These findings show that amifostine exhibits time dependent protection against cisplatin toxicity and thus it is recommended to use the protector when treatments are given during morning hours. The results also further validate the notion that chronochemotherapy is advantageous at least in reducing drug toxicity and thus should be integrated in the design of clinical protocols.

Prolactin rhythms-oscillators' response to photoperiodic cues is age and circadian time dependent.

In vivo prolactin release patterns exhibit a compound rhythm with circadian (24 h), semicircadian (12 h) and ultradian (6-8 h) periods. Changes in these rhythmic patterns were observed at different photoperiodic conditions, and in elderly. Since in vitro prolactin release was related to the photoperiodic history of the animal, we studied the effect of varying photoperioda upon the in vitro rhythmic output of prolactin release from young and old male rat pituitaries, isolated at different circadian times from animals housed at LD 12:12, 18:6 for 10 days or 6 weeks. The results indicate that, both, mean levels and rhythmic prolactin release in vitro are determined by the age of the animal, the circadian time of pituitary isolation, the photoperiodic conditions in which the animal was housed, and the duration of housing in the long day conditions. The change of the rhythmic output pattern is gradual, reflecting a process by which the oscillators respond to the external cues to fit prolactin release pattern to the environmental conditions. Each of the oscillators (e.g. circadian, semicircadian, ultradian) shows different sensitivity to the changing photoperiodic signal and is regulated at the level of phase and amplitude but not the period. In old rats the response of the oscillators to the change in photoperioda is attenuated and not sufficient to induce a change in the output of prolactin release suggesting a loss in adaptation ability.

Gender-dependent differences in biological rhythms of mice.

The advantage of a variable's rhythm resides in its optimal time-phasing. This implies that, for a given function, members of a species will strive to exhibit identical time-phasing namely, their inter-individual genetic differences will be masked. To examine the generality of this assumption we explored if inbred mice exhibit gender dependent differences in rhythm parameters of biochemical variables. Male and female mice, entrained by exposure to 12:12 light:dark illumination were sacrificed, every 3 hours over a 27 hours period. Activities of creatine-phosphokinase (CK) and alkaline- phosphatase (AP), white blood cell (WBC) counts and urea nitrogen (UN) concentration were determined at each time point. For each significant rhythm four parameters were computed: period, acrophase, mesor and amplitude. In addition two derived parameters were also calculated: relative-amplitude (RA) and the rate of change in RA (CRA) which provide information about the slope and width of the peak. Patterns of most variables exhibited a compound rhythm containing two significant periodicities. Gender dependent differences were documented in the parameters of most rhythms indicating that the genetic and physiological differences limit to a certain extent the phasing ability of the entraining signals and point to an independent control of each of the rhythm parameters.

Gonadotropin releasing hormone (GnRH) and estradiol (E(2)) regulation of cell cycle in gonadotrophs.

The number of pituitary cells, their size, hormonal content and release and response to external cues varies between day and night and during the estrus cycle. Previous studies have demonstrated that pituitary cells proliferate rhythmically and that estradiol (E(2)) is a mitogen of alpha T3 cells. We, therefore, studied the effect of gonadotropin releasing hormone (GnRH) and E(2), on the cell cycle in primary cultures of mouse pituitary cells and in the gonadotroph cell line L beta T2. We found that GnRH and E(2) modulate the cell cycle in a time dependent manner and induce proliferation in cultures of mouse pituitary and L beta T2 cells. GnRH induces proliferation in cells isolated in the morning of the estrus day and increases the number of cells in G2 stage when isolated in noon and evening. However, the transition into the G1 stage is enabled only by co-addition of E(2) and GnRH. GnRH stimulates LH release from L beta T2 cells after 2 days via exocytosis while after 4 days in culture, the increase in LH release may be accounted for by the increase in cell number. E(2) enhanced the GnRH response after 2 days, and abolished it after 4 days in culture. Furthermore, E(2) has no effect on LH release and cell number after 2 days in culture, however, after 4 days in culture, E(2) had no effect on the total amount of LH released but inhibited LH release per cell due to increase in cell number. Our results show that GnRH and E(2) function to shorten the cell cycle and regulate the cell number of each stage of the cell cycle. The effect of GnRH and E(2) on the cell cycle is dependent on the circadian time. This mechanism may serve to modulate the size and function of the pituitary cell population and consequently the function of pituitary gonadotrophs regulating the surge of LH release before ovulation.

Temporal pattern of LH secretion: regulation by multiple ultradian oscillators versus a single circadian oscillator.

The possibility that the 24h rhythm output is the composite expression of ultradian oscillators of varying periodicities was examined by assessing the effect of external continuously or pulsed (20-minute) Gonadotropin-releasing hormone (GnRH) infusions on in vitro luteinizing hormone (LH) release patterns from female mouse pituitaries during 38h study spans. Applying stepwise analyses (spectral, cosine fit, best-fit curve, and peak detection analyses) revealed the waveform shape of LH release output patterns over time is composed of several ultradian oscillations of different periods. The results further substantiated previous observations indicating the pituitary functions as an autonomous clock. The GnRH oscillator functions as a pulse generator and amplitude regulator, but it is not the oscillator that drives the ultradian LH release rhythms. At different stages of the estrus cycle, the effect of GnRH on the expression of ultradian periodicities varies, resulting in the modification of their amplitudes but not their periods. The functional output from the system of ultradian oscillators may superimpose a "circadian or infradian phenotype" on the observed secretion pattern. An "amplitude control" hypothesis is proposed: The temporal pattern of LH release is governed by several oscillators that function in conjunction with one another and are regulated by an amplitude-controlled mechanism. Simulated models show that such a mechanism results in better adaptive response to environmental requirements than does a single circadian oscillator.

Mechanism of GnRH receptor signaling on gonadotropin release and gene expression in pituitary gonadotrophs.

Gonadotropin releasing hormone (GnRH), the first key hormone of reproduction, is synthesized and secreted from the hypothalamus in a pulsatile manner and stimulates pituitary gonadotrophs (5-10% of the pituitary cells) to synthesize and release gonadotropin luteinizing hormone (LH) and follicle stimulating hormone (FSH). Gonadotrophs consist of 60% multihormonal cells (LH+FSH) and 18% LH- and 22% FSH-containing cells. LH and FSH, members of the glycoprotein hormone family, stimulate spermatogenesis, folliculogenesis, and ovulation. Although GnRH plays a pivotal role in gonadotropin synthesis and release, other factors such as gonadal steroids and gonadal peptides exert positive and negative feedback mechanisms, which affect GnRH actions. GnRH actions include activation of phosphoinositide turnover as well as phospholipase D and A2, mobilization and influx of Ca2+, activation of protein kinase C (PKC) and mitogen-activated protein kinase (MAPK). A complex crosstalk between the above messenger molecules mediates the diverse actions of GnRH. Understanding the signaling mechanisms involved in GnRH actions is the basis for our understanding of basic reproductive functions in general and gonadotropin synthesis and release in particular.

Circadian pattern of simulated flight performance of pilots is derived from ultradian components.

Studies suggest some physiologic, cognitive, and behavioral 24h rhythms are generated by cyclic components that are shorter in period than circadian. The aim of this study was (1) to examine the hypothesis that 24h human performance rhythms arise from the integration of high-frequency endogenous components and (2) to quantify the contribution of each higher frequency component to the phenotype of the rhythm. We monitored the performance of 9 experienced pilots by employing an array of cognitive-based tests conducted in a flight simulator so that, over the 6-day experiment, data were obtained for each 2h interval of the 24h. The activity-rest schedule of the subjects, no matter the exact clock time schedule of sleep and activity, always consisted of 14h activity (when they carried out regular professional duties) and 10h rest, with at least 8h of sleep. The simulated combat scenarios consisted of simple and complex tasks associated with target interception, aircraft maneuvering, and target shooting and downing. The results yielded two indices: the number of prominent periodicities in the time series and the relative magnitude of the amplitude of each relative to the construction of the composite 24h waveform. Three cyclic components (8h, 12h, and 24h) composed the observed 24h performance pattern. The dominant period and acrophase (peak time) of the compound output rhythm were determined by the interplay between the amplitudes of the various individual ultradian components. Task complexity (workload) increases the expression of the ultradian entities in the 24h pattern. We constructed a model composed of the multiple ultradian components; the composite output defined a "time span" (of 2h-4h duration) as opposed to an exact "time point" of high and low performance, endowing elevated functional capability.

From ultradian to infradian rhythms: LH release patterns in vitro.

In the present study, we examined in vitro luteinizing hormone (LH) release patterns from pituitaries and from pituitary cell cultures (3 and 7 days in culture) to elucidate the endogenous period generated by the gonadotroph cell population and to evaluate the relationship between the basic period generated at the cellular level and the output pattern observed at the organ level. In addition, we examined the effect of photic environmental signals perceived by the animals on LH release patterns from pituitaries in vitro. When the animals were exposed to circadian photoperiodic signals, the in vitro LH release pattern from the pituitaries exhibited ultradian, circadian, and infradian frequencies. When the animals were exposed to continuous illumination, the in vitro patterns exhibited only ultradian and infradian frequencies. Furthermore, free running is a process, not a state. This process is driven by a change in the relative dominance of different frequencies that construct the pattern without changing the basic period length. Evaluation of the relative dominance of the different frequencies that construct the pattern indicates that, although infradian oscillators may take part in shaping the output pattern, the basic rhythm generated by the pituitary cells is in the ultradian domain. The results obtained from the examined system suggest that an endogenous oscillator is a cellular entity with ultradian periodicity, and that the rhythmic output of many biological variables is structured by various ultradian components that construct the circadian and infradian output rhythms.

Mechanism of GnRH receptor signaling: from the membrane to the nucleus.

The purpose of this review is to update the information concerning the intracellular effect of GnRH. Binding of GnRH to a G-protein coupled receptor leads to stimulation of Gq and/or G11 protein and to activation of phospholipase C beta. Inositol 1-4-5-triphosphate and early diacylylycerol are the second messengers required for conventional protein kinase C activation. Activation of phospholipase A2 and phospholipase D are also involved, as demonstrated by the liberation of Arachidonic Acid and Phosphatidic Acid. Pituitary cells also express atypical protein kinase C isoforms which mode of activation is not known. Hypothesis concerning transcriptional regulation are presented.

Time dependency of hematopoietic growth factor coupled to chronotoxicity of carboplatin.

A growing body of data suggests that cancer therapy may be improved and toxicity reduced by administration of antineoplastic agents and cytokines at carefully selected times of the day. The time-dependent effects of each of the drugs have been documented, but not their mutual time dependencies. In the present studies we sought to determine the best time for granulocyte colony-stimulating factor (G-CSF) administration after carboplatin treatment. Carboplatin was injected in different groups of ICR mice at four different circadian stages for 5 consecutive days. Mice were synchronized with an alternation of 12 h of light (from 6:00 a.m. to 6:00 p.m.) and 12 h of darkness. After the last injection, peripheral WBCs of three mice from each group were counted every 4 h over a 24-h period. Bone marrow toxicity was estimated with the mean 24-h WBC count. The most severe leukopenia occurred in the group injected at 3:00 p.m. - 9 h after light onset. The second set of experiments evaluated the time-dependent effect of G-CSF when singly injected or given after carboplatin injections for 5 days only at 3:00 p.m. G-CSF was injected into various groups on days 8 and 9 at the same four different circadian stages. On the 10th day after the first injection, peripheral WBCs of three mice from each group were counted every 4 h over a 24-h period. Time-dependent effects were observed when G-CSF was injected as a single agent. When G-CSF was given at various times to the group with the most severe carboplatin-induced leukopenia, peripheral WBC count recovery was monitored at all injection times; it reached its highest level (exceeding even that of the control) when G-CSF was injected at 3:00 a.m. Dosing times of both chemotherapy and growth factor are relevant for optimization of carboplatin's hematologic tolerability.

The use of organizational principles in naming generation: its relation to time-dependency in normal elderly and Alzheimer patients.

Nineteen elderly--ten healthy and nine Alzheimer patients--were given a naming-generation task. Their responses were analyzed according to word-type and inter-word relatedness. The results were correlated with the Mini-Mental-State Examination (MMSE) and examined to elucidate time-dependency. In healthy subjects it was found that time-dependency exists for the number of new words and for percent of phonetically-related words, while for Alzheimer patients, time-dependency was exhibited in new words only. Results are discussed in terms of the principles of semantic-memory organization and time-dependency of cognitive-linguistic functions.

Interindividual differences in the flexibility of human temporal organization: pertinence to jet lag and shiftwork.

Interindividual variability in the human temporal structure is seldom taken into account, especially in studies devoted to the effects of shiftwork and jet lag. The understated postulate is that humans can be treated as a pure strain species. This paper reviews some facts and concepts with special reference to interindividual changes in the rhythm period tau and the resulting dyschronism. The following points are addressed. (1) Subjects and methods (importance of longitudinal field studies on shift workers). (2) Criteria for tolerance to shiftwork and jet lag. (3) Interindividual differences and shiftwork problems (subject type; the association between good shiftwork tolerance and stable temporal structure; dychronism with tau s differing from 24h and from variable to variable. (4) The genetic background of circadian dyschronism. The Dian-circadian genetic model of biological rhythms. It allows understanding of one's susceptibility to dyschronism, which was actually observed in approximately equal to 30% of subjects studied longitudinally. (5) Practical implications of interindividual differences (dissociate problems of passengers after a transmeridian flight-who have to adjust their temporal structure to local time-from problems of shiftworkers-who need to prevent alteration of their temporal structure; the advantage for the latter of participating in a rapid rotation system rather than a weekly rotation; emphasis that the suitability of a given subject for a given shiftworking condition is likely to be estimated only after a trial span of time including longitudinal study of a set of rhythms.

Differences between left- and right-hand reaction time rhythms: indications of shifts in strategies of human brain activity.

Reaction time (RT) measurements serve as quantitative indices for pilots' cognitive processes of the brain. To examine if laterality exists in the brain hemispheres we measured, by the use of a Pilot Evaluation System (PES), right- and left-hand performance rhythms as indicative of RT to audible and visual stimuli. The tests included sets of simple tasks and complex ones to which a secondary task composed of audio signals was added. The accuracy of recorded reaction time was 27 ms. Seven right-handed males, 27-42 years of age, experienced with the PES flight simulator, were tested every 2 h, nine times daily (starting at 08:00 h) during 3 consecutive days. The results indicated that for simple tasks, the 24 h period of RT rhythm is either exclusive or prominent for both hands. For complex tasks the prominent period of RT is 24 h for the right (dominant) hand and 8 h for the left (non-dominant) hand (right-hand 24 h period Fstat = 140, r2 = 0.62 and 8 h period Fstat = 25, r2 = 0.22; left-hand 24 h period Fstat = 44, r2 = 0.34 and 8 h period Fstat = 100, r2 = 0.54). The findings suggest that a laterality exists in the brain hemispheres with regard to differences in rhythm periodicities. The expression of this laterality is dependent on the task-load level and points to a strategy of linkage and integrity in brain activity.

Two types of growth hormone rhythm in boys with constitutional short stature.

The relationship between temporal and quantitative changes in pulsatile growth hormone (GH) secretion and growth of constitutionally short children (CSS) was studied in 19 prepubertal boys and 4 with early adrenarche aged 7.8 to 14 years. Continuous plasma GH monitoring was performed at 30 minutes intervals during 24 hours. The data were analysed by single and serial best fit (BFT) cosinor analysis for rhythm parameters and the Pulsar peak detection program for determining pulsatile properties. The pulsatile patterns were expressed in mean GH concentration, area under the curve, mean pulse area and maximal pulse height but not in the number of pulses. Thirteen out of 23 patterns exhibited significant 24-h compatible rhythms. The studied cohort was divided into two groups, those who exhibited the 24-h circadian rhythm (CIR) and those who lacked it (NCIR). Ultradian 2.5-4 h rhythms were observed in all patients. The NCIR children were significantly shorter than the CIR group (p = 0.017). The CIR boys secreted a significantly higher (p = 0.01) amount of GH during the 24-h span, most of which was during sleep. Our data indicate that the magnitude of pulses rather than their number is responsible for the quantitative differences observed between the two groups of boys with CSS. It is suggested that a lack of 24-h rhythmicity and the associated reduced nocturnal pulsatility play a role in the manifestation of short stature.

Rhythmicity of luteinizing hormone secretion expressed in vitro.

In the present study we explored the possibility that the pituitary functions as an autonomous clock and is capable of generating rhythms of luteinizing hormone (LH) release independently of hypothalamic control. Pituitaries from estrous or diestrous day 1 female mice were perifused separately with Medium-199. Effluent samples were collected at 10-min intervals and assayed for LH levels. Fourier analysis and curve-fit analysis served to elucidate the presence of prominent periods whose significance was then determined by best-fit cosinor. The latter method was used to determine additional parameters for the significant rhythm. All perifused pituitaries exhibited LH release patterns that were composed of significantly long ultradian rhythms (approximately 16 and 8 h, p < 0.001). Continuous stimulation with gonadotropin-releasing hormone (GnRH) or estradiol did not alter the periods of the observed rhythms but affected other rhythm parameters. Gonadotropin-releasing hormone increased the mesor of the rhythm and estradiol increased the amplitude. The results indicate that pituitary gonadotropes are capable of producing rhythms of LH release for a long duration in vitro, in the absence of hypothalamic control. Both GnRH and estradiol affect different rhythm parameters but do not change the periods of these rhythms.

The pattern of hormonal circadian time structure (acrophase) as an assessor of breast-cancer risk.

Through many hormones are secreted in a pulsatile manner, their secretion pattern can be superimposed by a 24-hour sinusoidal curve. The sinusoidal curve is then characterized by the estimated peak clock time location (acrophase), the adjusted mean (mesor) and the amplitude. When the distribution of the acrophases of 12 hormones was compared among women with regard to their age and to the level of risk of developing breast cancer, statistically significant differences were revealed between distribution patterns of acrophases of women with high (n = 12 and 45 circadian profiles) or low (n = 12 and 41 circadian profiles) risk of developing breast cancer. However, when the amplitude/mesor ratios of the corresponding hormonal rhythms were analyzed, significant differences occurred between age groups rather than between risk levels. These observations suggest that the endocrine time structure between individual women can be used as an assessor of breast-cancer risk.

Prevalence of genetic versus environmental factors in human female temporal organization: preliminary analysis.

Genetic diversity among ethnic groups is studied by comparing the genetic fingerprint of the examined groups. This index is constructed by aggregating the differential frequencies of various marker characteristics. Recent advances in the study of human biological rhythms may provide new indexes that will complement the genetic profile of a population. One of the rhythm parameters that is especially useful for this purpose is the acrophase (peak time location). The aim of the present study is to construct a rhythm profile based on acrophase distribution for various human groups and to estimate the contribution of genetic and environmental factors to that profile. The rhythm profiles were constructed by comparing the acrophases of 11 plasma hormones in women from three different ethnic-geographic populations (North Americans, Romanians, and Japanese) with reference to three age groups (adolescence-early postpuberty, young adulthood, and postmenopause). Genetic distances of these ethnic groups were determined by 14 genetic markers. Cluster and principal coordinates analyses were used to define the variation of the two parameters (genetic distances and acrophase dispersion). The analyses show that North Americans and Romanians are closer to each other with regard to both parameters and far apart from the Japanese. However, there was a difference between the variation presented by the first eigenvalue of the genetic profiles (94.5%) and that of the first eigenvalue of the acrophase pattern (69.1%), which means reduction in the variability (increased similarity) among the three ethnic groups according to the acrophase profiles.(ABSTRACT TRUNCATED AT 250 WORDS)

Superactive lipophilic peptides discriminate multiple vasoactive intestinal peptide receptors.

To distinguish vasoactive intestinal peptide (VIP) receptors in the brain-mediating neurotransmission and neurotrophism, potent VIP analogues were designed. Using a single amino acid substitution and the addition of a fatty acyl moiety, an analogue was devised that exhibited both a 100-fold greater potency than VIP and specificity for a VIP receptor associated with neuronal survival. This VIP agonist increased neuronal survival via a cAMP-independent mechanism. Identical chemical modification of a prototype VIP antagonist (Met-Hybrid, Neurotensin6-11-VIP7-28) also resulted in a 100-fold greater potency in blocking VIP-mediated increases in neuronal survival. Blockade of circadian activity rhythms was limited to VIP antagonists that could inhibit VIP-mediated increases in cAMP. These lipophilic peptides provide novel tools in receptor discrimination and drug design.