Body composition and metabolic changes during a 520-day mission simulation to Mars.

PURPOSE: The "Mars-500 project" allowed to evaluate the changes in psychological/physiological adaptation over a prolonged confinement, in order to gather information for future missions. Here, we evaluated the impact of confinement and isolation on body composition, glucose metabolism/insulin resistance and adipokine levels. METHODS: The "Mars-500 project" consisted of 520 consecutive days of confinement from June 3, 2010 to Nov 4, 2011. The crew was composed of six male subjects (three Russians, two Europeans, and one Chinese) with a median age of 31 years (range 27-38 years). RESULTS: During the 520-day confinement, total body mass and BMI progressively decreased, reaching a significant difference at the end (417 days) of the observation period (- 9.2 and - 5.5%, respectively). Fat mass remained unchanged. A progressive and significant increase of fasting plasma glucose was observed between 249 and 417 days (+ 10/+ 17% vs baseline), with a further increase at the end of confinement (up to + 30%). Median plasma insulin showed a non-significant early increment (60 days; + 86%). Total adiponectin halved (- 47%) 60 days after hatch closure, remaining at this nadir (- 51%) level for a further 60 days. High molecular weight adiponectin remained significantly lower from 60 to 168 days. CONCLUSIONS: Based on these data, countermeasures may be envisioned to balance the potentially harmful effects of prolonged confinement, including a better exercise program, with accurate monitoring of (1) the individual activity and (2) the relationship between body composition and metabolic derangement.

Diagnostic accuracy of 1.5 Tesla breast magnetic resonance imaging in the pre-operative assessment of axillary lymph nodes.

The purpose of this study was to test the accuracy of 1.5 Tesla magnetic resonance imaging (1.5T MRI) in the preoperative evaluation of axillary lymph nodes in patients with invasive breast cancer. The authors retrospectively analyzed 26 patients with invasive breast cancer who had undergone sentinel lymph node biopsy (SLNB) and/or axillary lymph node dissection (ALND). All patients had been submitted to preoperative contrast enhanced breast 1.5T MRI. On the basis of lymph nodes morphological and dynamic characteristics, lymph nodes were classified as "negative" (short axis < 5 mm), "borderline" (short axis > 5 mm, absence of a hilum) or "positive" (short axis > 5 mm, absence of a hilum and also other suspicious features). The authors compared 1.5T MRI results with the outcome of histological analysis performed according to the TNM criteria; sensitivity (SE), specificity (SP), positive predictive value (PPV), and negative predictive value (NPV) of 1.5T MRI were evaluated. Considering only the lymph nodes "positive", 1.5 T MRI showed: SE 37.8%, SP 99.3%, FP 0.7%, PPV 92.5%, and NPV 88.1%. However, considering also "borderline", 1.5T MRI achieved: SE 75.7%, SP 99.3%, FP 0.7%, PPV 96.1%, and NPV was 95%. Contrast enhanced breast 1.5T MRI is not yet a valid alternative to histological analysis but it is a valid tool for a preoperative study of the topography of axillary lymph nodes and has the potential to become a routine method for evaluating the metastatic lymph nodes before submission to ALND.

Apparent diffusion coefficient on 3.0 Tesla magnetic resonance imaging and prognostic factors in breast cancer.

AIM: The aim of the study was to evaluate whether the apparent diffusion coefficient (ADC) provided by 3.0 Tesla diffusion-weighted imaging (3T DWI) varies with the prognostic factors Ki67 and grading in invasive breast cancer. MATERIALS AND METHODS: Seventy-three patients with 75 invasive breast cancer lesions who had undergone 3.0 Tesla magnetic resonance imaging (MRI) for local staging were enrolled. All lesions were confirmed by histologic and immunohistochemical analysis. MRI included both dynamic contrast-enhanced and DWI sequences. ADC value was obtained for each lesion. Histologic tumor grade was established according to the Nottingham Grading System (NGS), while Ki67 expression was evaluated by MM1 clone IgG1 mouse anti-human monoclonal antibody. Patients were divided into the following groups: grade 1 (G1), grade 2 (G2), grade 1 plus grade 2 (G1+G2) and grade 3 (G3), and low Ki67 (< or = 14%), intermediate Ki67 (15%-30%), and high Ki67 (> or = 30%). ADC values were compared with the G and Ki67 groups. Statistical comparison was carried out using the Mann-Whitney U and the Kruskal-Wallis H test. RESULTS: ADC values were significantly higher in G3 than in G1+G2 tumors; no significant difference was observed when G1, G2, and G3 were compared. There was no statistically significant correlation between ADC values and Ki67 percentage (p > 0.05). DISCUSSION: ADC values obtained on 3T DWI correlate with low (G1+G2) and high-grade (G3) invasive breast carcinomas. CONCLUSION: ADC may be a helpful tool for identifying high-grade invasive breast carcinoma.

Role of psa levels and pathological stadiation before radiation therapy in predicting mp-MRI results in patients with prostate cancer recurrence after radical prostatectomy.

OBJECTIVE: To evaluate PSA value in mp-MRI results prediction, analyzing patients with high (GS≥8, pT≥3, pN1) and low grade (GS<8, pT<3, pN0) Prostate Cancer (PCa). MATERIALS AND METHODS: One hundred eighty-eight patients underwent 1.5-Tmp-MRI after Radical Prostatectomy (RP) and before Radiotherapy (RT). They were divided into 2 groups: A and B, for patients with biochemical recurrence (BCR) and without BCR but with high local recurrence risk. Considering Gleason Score (GS), pT and pN as independent grouping variables, ROC analyses of PSA levels at primary PCa diagnosis and PSA before RT were performed in order to identify the optimal cut-off to predict mp-MRI result. RESULTS: Group A and B showed higher AUC for PSA before RT than PSA at PCa diagnosis, in low and high grade tumors. For low grade tumors the best AUC was 0.646 and 0.685 in group A and B; for high grade the best AUC was 0.705 and 1 in group A and B, respectively. For low grade tumors the best PSA cut-off was 0.565-0.58ng/mL in group A (sensitivity, specificity: 70.5%, 66%), and 0.11-0.13ng/mL in B (sensitivity, specificity: 62.5%, 84.6%). For high grade tumors, the best PSA cut-off obtained was 0.265-0.305ng/mL in group A (sensitivity, specificity: 95%, 42.1%), and 0.13-0.15ng/mL in B (sensitivity, specificity: 100%). CONCLUSION: Mp-MRI should be performed as added diagnostic tool always when a BCR is detected, especially in high grade PCa. In patients without BCR, mp-MRI results, although poorly related to pathological stadiation, still have a good diagnostic performance, mostly when PSA>0.1-0.15ng/mL.

Impact of adipose tissue distribution on prostate cancer recurrence after radical prostatectomy.

OBJECTIVE: The aim of our study is to correlate the CT adipose tissue distribution and recurrence risk of Prostatic Cancer (PCa) after Radical Prostatectomy (RP). Furthermore, we evaluated the association of adipose tissue and PCa aggressiveness. MATERIALS AND METHODS: We identified two groups of patients based on presence (group A) and absence (group B or control group) of Bio-chemical Recurrence (BCR) after RP. A semi-automatic function able to recognize the typical attenuation values of adipose tissue was used for sub-cutaneous (SCAT), visceral (VAT), total (TAT) and periprostatic (PPAT) adipose tissues. For both groups of patients, a descriptive analysis of continuous variables and categorical variables was performed. RESULTS: After comparing between groups, a statistically significant difference was found for VAT (p<0.001) and for VAT/TAT ratio (p=0.013). No statistically significant correlation was found for PPAT and SCAT, even if higher values were found in patients with high grade tumors. CONCLUSION: This study confirms visceral adipose tissue as a quantitative imaging parameter related to oncological risk of PCa recurrence development, and the role of abdominal fat distribution measured with CT before RP as an important tool to predict the PCa recurrence risk, particularly in patients with high grade tumors.

Endovascular radiofrequency renal denervation in treating refractory arterial hypertension: a preliminary experience.

PURPOSE: This study was done to investigate the efficacy and safety of percutaneous renal denervation with the Symplicity catheter for reducing blood pressure in patients with essential hypertension resistant to medical therapy (systolic blood pressure >160 mmHg despite the use of three or more antihypertensive drugs, including a diuretic). MATERIALS AND METHODS: In September 2010, five patients affected by essential hypertension resistant to medical therapy were treated. All patients were studied by computed tomography angiography (CTA) of the renal arteries before the procedure and underwent follow-up at 30 and 60 days with colour Doppler ultrasound (CDUS) with evaluation of resistive index, glomerular filtration rate (GFR), 24-h blood pressure and serum catecholamine concentration. Student's t test was used to assess the effectiveness of the procedure in lowering blood pressure. RESULTS: In treated patients, mean blood pressure at baseline was 171/100 mmHg [standard deviation (SD) ± 8/10]; mean GFR was 91.6 ml/min/1.73 m(2) (SD ± 15). Blood pressure after the procedure was reduced by -18/-5 and -13/-10 mmHg at 30 and 60 days, respectively, with a mean medication reduction of 3.6. No complications occurred during the intra- or periprocedural period or during short-term follow-up. CONCLUSIONS: The Symplicity system proved to be efficacious and without serious adverse events in reducing blood pressure and antihypertensive medication use in patients affected by essential hypertension resistant to medical therapy. Although encouraging, our data are preliminary and need to be validated by larger prospective randomised studies.

The role of multiparametric mri in the diagnosis of local recurrence after radical prostatectomy and before salvage radiotherapy.

PURPOSE: Assess multiparametric-MRI (mp-MRI) diagnostic accuracy in the detection of local recurrence of Prostate Cancer (PCa) after Radical Prostatectomy (PR) and before Radiation Therapy (RT). MATERIALS AND METHODS: A total of 188 patients underwent 1.5-T mp-MRI after RP before RT. Patients were divided into two groups: with biochemical recurrence (group A) and without but with high risk of local recurrence (group B). Continuous variables were compared between two groups using T-Student; categoric variables were analyzed using Pearson chi-square. ROC analysis was performed considering PSA before RT, ISUP, pT and pN as grouping variables. RESULTS: PCa recurrence (reduction of PSA levels after RT) was 89.8% in the group A and 80.3% in the group B. Comparing patients with and without PCa recurrence, there was a significant difference in PSA values before RT for group A and for PSA values before RT and after RT for group B. In group A, there was a significant correlation between PSA before RT and diameter of recurrence and between PSA before RT and time spent before recurrence. The mp-MRI diagnostic accuracy in detecting PCa local recurrence after RP is of 62.2% in group A and 38% in group B. DWI is the most specific MRI-sequence and DCE the most sensitive. For PSA = 0.5 ng/ml, the AUC decreases while sensitivity and accuracy increase for each MRI-sequence. For PSA = 0.9 ng/ml, DCE-AUC increases significantly. CONCLUSION: mp-MRI should always be performed before RT when a recurrence is suspected. New scenarios can be opened considering the role of DWI for PSA ≤ 0.5 ng/ml.

The endocrine disruptor atrazine accounts for a dimorphic somatostatinergic neuronal expression pattern in mice.

It has now been established that a large number of man-made and natural chemicals are capable of interfering with the action of natural hormones. In this category "endocrine disruptors" such as the herbicide atrazine, when administered at ecological low doses (1 or 100 microg/kg per day) from gestational day 14 to postnatal day 21, provided a clear dimorphic neurodegenerative pattern in some brain areas of the domestic mouse (Mus musculus). Indeed, the high concentration (100 microg/kg per day) with respect to the low concentration (1 microg/kg per day) induced relevant neuronal damage in extrahypothalamic sites, such as the cortical and striatal areas in both sexes. Marked alterations in other areas, including the hippocampal and hypothalamic nuclei, were mostly typical of the female. At the neuronal level, the neuropeptide somatostatin, specific for the secretion of growth hormone, seemed to be a major target of atrazine effects, as demonstrated by evident subtype2,3,5 receptor mRNA differences of this neuropeptide, at least for the first two subtypes. In particular, a very strong (p < 0.001) upregulation of subtype2 expressing neurons was detected in female hypothalamic areas, specifically the suprachiasmatic nucleus, whereas a similar downregulatory trend was reported for some extrahypothalamic areas such as the striatum. Interestingly, very strong upregulatory and downregulatory actions were detected for neurons expressing subtype3 in male hypothalamic and amygdalar regions and in the cortical and hippocampal areas, respectively. Overall, it appears that these first neurotoxicological effects of atrazine are very likely linked to dimorphic expression patterns of specific somatostatin subtypes in discrete but key hypothalamic and extrahypothalamic areas of Mus musculus.

Regulation of the inducible soybean nitrate reductase isoform in mutants lacking constitutive isoform(s).

In soybean, three nitrate reductase isoforms have been identified based on metabolic regulation, substrate specificity, and kinetic parameters. Two isoforms have been termed constitutive, as their activities are present without the addition exogenous nitrate to soybean seedlings. The third activity is termed inducible, as its activity is present only when soybean plants have been supplied with nitrate. The purpose of this study was to examine the regulation of the inducible nitrate reductase isoform in soybean mutants lacking one or both of the constitutive isoforms. Based on evidence obtained through measurements of enzyme activity, Western blotting, and RNA determinations, the absence of one or both of the constitutive nitrate reductase isoforms has no effect on the metabolite regulation of the inducible nitrate reductase isoform.

Neurobehavioural effects of hypergravity conditions in the adult mouse.

To evaluate the behavioural response to a hypergravity condition in CD-1 mice, young adult subjects of both sexes were exposed to 2 g for a single 60 min rotational session. Motion sickness (MS) and ethological-type scoring of different activities were used to evaluate the behavioural response. Nerve growth factor (NGF) and brain derived neurotrophic factor (BDNF) levels were also assessed. Behavioural scores indicated a transient mild sickness associated with hypergravity, with reduction in spontaneous activity. In males kaolin consumption (a MS index) increased following rotation while females consumed more kaolin irrespective of whether they have been rotated or simply exposed to the noise and vibration of the rotational apparatus. In males, hypothalamic NGF levels were markedly increased after rotation while no major changes were observed in central BDNF expression. These results indicate mice may represent a suitable MS model.

Ultrasonic vocalisation emitted by infant rodents: a tool for assessment of neurobehavioural development.

Ultrasonic vocalisations (USVs) emitted by altricial rodent pups are whistle-like sounds with frequencies between 30 and 90 kHz. These signals play an important communicative role in mother-offspring interaction since they elicit in the dam a prompt response concerning caregiving behaviours. Both physical and social parameters modulate the USV emission in the infant rodent. Recently, a more detailed analysis of the ultrasonic vocalisation pattern, considering the spectrographic structure of sounds has allowed a deeper investigation of this behaviour. In order to investigate neurobehavioural development, the analysis of USVs presents several advantages, mainly: (i) USVs are one of the few responses produced by very young mice that can be quantitatively analysed and elicited by quantifiable stimuli; (ii) USV production follows a clear ontogenetic profile from birth to PND 14-15, thus allowing longitudinal neurobehavioural analysis during very early postnatal ontogeny. The study of this ethologically-ecologically relevant behaviour represent a valid model to evaluate possible alterations in the neurobehavioural development of perinatally treated or genetically modified infant rodents. Furthermore, the role played by several receptor agonists and antagonists in modulating USV rate makes this measure particularly important when investigating the effects of anxiogenic and anxiolytic compounds, and emotional behaviour in general.

Early exposure to aluminium affects eight-arm maze performance and hippocampal nerve growth factor levels in adult mice.

Aluminium has been implicated in the etiology of several human pathologies but, to date, little attention has been given to the potential damage caused by gestational exposure to aluminium or to any long-term effect of early exposure. Aluminium sulphate (200 mg/kg) was administered intraperitoneally to pregnant female mice (days 10 to 13 of gestation), and male progeny was assessed in a radial eight-arm maze task at 70 days (20 daily sessions, 50 microliters water as reward). Overall, aluminium-exposed mice performed less efficiently in the radial maze than the vehicle-exposed animals. Neurobiological data showed that aluminium mice also had a higher level of nerve growth factor in the hippocampal formation when compared to controls.

Effect of hypergravity on the mouse basal expression of NGF and BDNF in the retina, visual cortex and geniculate nucleus: correlative aspects with NPY immunoreactivity.

We investigated the effect of hypergravitation on Nerve growth factor (NGF) and Brain-derived-neurotrophic factor (BDNF) expression in the visual cortex, geniculate nucleus (GN), and retina of adult male mice. The results showed that altered gravity causes an increase in NGF and BDNF in the visual cortex and GN which resulted to be associated with an up-regulation of cells immunoreactive to neuropeptide Y (NPY) in the visual cortex and GN. We also found a decrease in NGF, BDNF, and NPY in the mouse retina exposed to hypergravity. These findings suggest that alteration in gravitational environment differentially affects local neurotrophic factors and NPY expression. The possible functional significance of these observations is discussed.

IGF-I and IGF-I24-41 but not IGF-I57-70 affect somatic and neurobehavioral development of newborn male mice.

Insulin-like growth factor (IGF-I) is a trophic factor for both neurons and glial cells. Its presence in developing and adult nervous system suggests an important role for this peptide in the development of neural circuitries. Neonatal male mice of the CD-1 outbred strain were injected intracerebroventricularly with either recombinant IGF-I, synthetic IGF-I fragment 24-41 or IGF-I fragment 57-70 on postnatal days (PND) 2, 4, and 7. Physical traits such as body weight gain, body length, and tail length were recorded daily from PND2 to PND13. Sensorimotor development was scored according to a modified Fox's scale. The ultrasonic vocalization pattern on PND8 and homing performance on PND10 were also recorded. Measures for body weight gain and tail length of the pups were significantly increased following treatment with the whole IGF-I peptide. However, neither IGF-I nor the smaller fragments affected mice sensorimotor development. IGF-I and IGF-I24-41 but not IGF-I57-70 increased the rate of ultrasonic calls of the pups measured on PND8. These data provide evidence that IGF-I regulates somatic growth and behavioral development when administered in newborn mice and that different portions of the peptide can exert different effects.

Effects of acute and continuous ozone (O3) exposure on activity/exploration and social behavior of CD-1 mice.

This study was aimed at investigating the behavioral effects of ozone (O3) exposure in CD-1 mice. Pairs of same-sex adult male and female mice were continuously exposed for 13 days to either 0, 0.4, 0.8, or 1.2 ppm O3. The exposure apparatus consisted of a system for O3 production and delivery into four stainless-steel chambers, each equipped to contain up to 24 home cages, with continuous monitoring and recording of concentrations. Acute behavioral changes were assessed during the first hour of O3 exposure without removing animals from the chambers. The onset of exposure produced remarkable behavioral disturbances consisting of a sharp increase of several responses (rearing, sniffing, grooming, feeding, and social interactions) paralleled by a reduction of bar-holding. These changes were rapidly reversed within 1 hour, suggesting that they constituted a response to strong novel stimulation followed by habituation. Subsequently, brief sessions of videorecording of the animals' activities in freshly cleaned cages (identical to the home cages) were performed outside the chambers after 3, 7, and 10 days of exposure. These tests showed a significant concentration-dependent increase of grooming and rearing and a decrease of crossing and wall climbing. Both food and water intake showed a nonmonotonic trend over time consisting of a concentration-dependent depression (for about 3 and 7 days, respectively) followed by recovery; body weight followed a similar trend. The detailed study of various components of the animal's behavioral repertoire, showing concentration-dependent and time-dependent changes in different directions, appears to be a sensitive tool in the analysis of pollutants' effects.

Psychosocial vs. "physical" stress situations in rodents and humans: role of neurotrophins.

Nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) are well-studied polypeptide growth factors involved in the development and maintenance of specific peripheral and central populations of neuronal cells. In addition to its role as a neurotrophic agent, NGF controls very complex functions in vertebrate physiology. A variety of cells outside the nervous system are in fact able to synthesize NGF including epithelial cells, fibroblasts, lymphocytes, and macrophages. NGF target cells have been identified in the nervous, immune, and endocrine systems, suggesting that NGF may operate through multiple paths to ultimately regulate physiological homeostasis and behavioral coping. We used a mouse model of social stress to demonstrate that NGF levels increase both in plasma and in the hypothalamus following intermale aggressive interactions. The investigation has been extended to other species, including humans, to show that labour, lactation, and the anticipation of the first jump with a parachute also result in increased NGF plasma levels and in changes in the distribution of NGF receptors on lymphocytes. BDNF activation is caused by both physical and social stress events. The aim of this review is to (1) outline the current understanding of the roles of NGF and BDNF in stress-related physiological changes in vertebrates, in particular for physical vs. psychological stressors, which may activate both similar and different neurobiological pathways, and (2) summarize recent efforts to derive pharmacological strategies from the increasing body of BDNF and NGF neurobehavioral data.

Response of CD-1 mice to the chemical defence of a common arthropod (Ommatoiulus sabulosus).

In order to set up a novel and ethologically relevant methodology that could be applied to the study of olfactory capabilities in transgenic mice, we analysed the behavioural responses of sexually mature male and female CD-1 mice individually exposed to a striped millipede, Ommatoiulus sabulosus (L.), a very common myriapod species that secretes a repulsive and persistent odour in the presence of a predator. As control, we exposed mice to a larva of the lepidopteran Greater wax moth, Galleria mellonella (L.), which closely resembles the millipede in shape and dimensions but which does not secrete a repulsive odour in defence. We recorded and analysed a wide spectrum of behavioural responses including both those of avoidance and nonavoidance such as attempts to eat the arthropod. Behavioural responses were measured for 10 min upon first exposure to the millipede or wax moth. The procedure was repeated for 3 consecutive days. Upon exposure to a millipede, mice of both sexes showed a dramatic increase in the avoidance behaviour of digging. Moreover, millipedes were repulsive to mice and though they were sniffed frequently and sometimes caught, they were never eaten. In comparison, mice exposed to a wax moth almost always ate it. Sex differences emerged only for locomotion with female appearing to be more active. These results suggest that mice are able to discriminate between ethologically relevant odours and that the behavioural responses they display in this more natural context differ from those observed in response to odours of predators.

Neonatal exposure to bFGF exerts NGF-like effects on mouse behavioral development.

Brain cells are naturally exposed to a variety of trophic factors during development. Basic Fibroblast Growth Factor (bFGF), a protein found in the central nervous system (CNS) enhances both survival and proliferation of several CNS cell lines. Neonatal mice of the CD1 outbred strain were injected intracerebroventricularly (ICV) with bovine bFGF on postnatal days 2, 4, and 7. bFGF, dissolved in 25 microliters of saline, was given in the 1, 5, 25, and 1000 ng doses. Physical traits such as body weight gain, body length, and tail length were recorded from postnatal day (PND) 2 to PND 12. Neurobehavioral development was scored according to a modified Fox's Scale. The ultrasonic vocalization pattern (PND 8), homing performance (PND 10), open-field activity and exploration of a novel object (PND 18) were also assessed. Pups receiving 25 and 1000 ng doses of bFGF showed a slight but significant acceleration in the appearance of some early sensorimotor reflexes, such as weak and tactile stimulation, forelimb placing and grasping, and screen climbing, during the first postnatal weeks. The other responses were less affected or not affected at all. Data are compared with similar systemic exposure of neonatal mice to Nerve Growth Factor (NGF).

Behavioural effects of endocrine disrupting chemicals on laboratory rodents: statistical methodologies and an application concerning developmental PCB exposure.

Appropriate behavioural tests and adequate statistical tools may help to establish the ED properties of a given compound by pointing out the alterations of selected behavioural endpoints. Frequently, laboratory collected data consist of frequencies and/or durations of specific items, and the analysis of variance (ANOVA) technique is performed to assess whether the investigated factors affect these behavioural endpoints. Moreover, when numerous aspects of behaviour are investigated simultaneously, Principal Component Analysis (PCA), a multivariate technique, may be very useful to reduce the overwhelming number of correlated original variables to a few orthogonal artificial variables (factors). Continuous Time Markov Chain (CTMC) models may be applied to analyse the time structure of a behavioural pattern when data consist of sequences of events and the time points at which they occur. Moreover, the Cox Proportional Hazard Model, a methodology originally developed for the analysis of failure time data, may help to evidence the effects of a given treatment on behavioural sequences when the assumptions of CTMC models are not fully satisfied. Analyses on data from mice of the outbred CD-1 strain (controls in a study of toxicity and exposed to PCB during development) are presented as examples to show how adequate statistical analyses and appropriate behavioural tests may reveal relevant effect of treatments otherwise not easily detected.

Effects of age, sex, and odours from conspecific adult males on ultrasonic vocalizations of infant CS1 mice.

Young CS1 mice aged 2, 4, 6, 8 and 10 days were each sequentially exposed to odours of urine from strange adult infanticidal and non-infanticidal males and the number of ultrasonic vocalizations monitored. There was a clear age effect, with pups 6-8 days calling the most. For pups of 2 and 6 days of age there were significant positive correlations between body weight and number of calls but for 10-day-old pups a negative correlation was found. Female pups called more than males, particularly when exposed to odours but this was not due to body weight. Overall, more calls were produced when odours from infanticidal rather than non-infanticidal males were presented and individual pups altered their ultrasonic output as the odours were changed.