Oxytocin (OT) is a neurosecretory nonapeptide synthesized in hypothalamic cells that project to the neurohypophysis as well as to widely distributed sites in the central nervous system. Central OT microinjections induce a variety of cognitive, sexual, reproductive, grooming and affiliative behaviors in animals. Obsessive-compulsive disorder (OCD) includes a range of cognitive and behavioral symptoms that bear some relationship with OT. Here, we study the neuroanatomical and cellular substrates of the hypergrooming induced by administration of OT in the central nucleus of amygdala (CeA). In this context, this hypergrooming is considered as a model of compulsive behavior. Our data suggest a link between the CeA and the hypothalamic grooming area (HGA). The HGA includes parts of the paraventricular nucleus and the dorsal hypothalamic area. Our data on colocalization of OT (immunohistochemistry for peptide), OT receptor (binding assay) and its retrogradely labeled cells after Fluoro-Gold injection in the CeA suggest that CeA and connections are important substrates of the circuit underlying this OT-dependent compulsive behavioral pattern.

There are reports of patients whose epileptic seizures are prevented by means of olfactory stimulation. Similar findings were described in animal models of epilepsy, such as the electrical kindling of amygdala, where olfactory stimulation with toluene (TOL) suppressed seizures in most rats, even when the stimuli were 20% above the threshold to evoke seizures in already kindled animals. The Wistar Audiogenic Rat (WAR) strain is a model of tonic-clonic seizures induced by acute acoustic stimulation, although it also expresses limbic seizures when repeated acoustic stimulation occurs - a process known as audiogenic kindling (AK). The aim of this study was to evaluate whether or not the olfactory stimulation with TOL would interfere on the behavioral expression of brainstem (acute) and limbic (chronic) seizures in the WAR strain. For this, animals were exposed to TOL or saline (SAL) and subsequently exposed to acoustic stimulation in two conditions that generated: I) acute audiogenic seizures (only one acoustic stimulus, without previous seizure experience before of the odor test) and II) after AK (20 acoustic stimuli [2 daily] before of the protocol test). We observed a decrease in the seizure severity index of animals exposed only to TOL in both conditions, with TOL presented 20s before the acoustic stimulation in both protocols. These findings were confirmed by behavioral sequential analysis (neuroethology), which clearly indicated an exacerbation of clusters of specific behaviors such as exploration and grooming (self-cleaning), as well as significant decrease in the expression of brainstem and limbic seizures in response to TOL. Thus, these data demonstrate that TOL, a strong olfactory stimulus, has anticonvulsant properties, detected by the decrease of acute and AK seizures in WARs.

IntroductionThe central nucleus of amygdala plays an important role mediating fear and anxiety responses. It is known that oxytocin microinjections into the central nucleus of amygdala induce hypergrooming, an experimental model of compulsive behavior. We evaluated the behavioral and cardiorespiratory responses of conscious rats microinjected with oxytocin into the central nucleus of amygdala.MethodsMale Wistar rats were implanted with guide cannulae into the central nucleus of amygdala and microinjected with oxytocin (0.5 µg, 1 µg) or saline. After 24 h, rats had a catheter implanted into the femoral artery for pulsatile arterial pressure measurement. The pulsatile arterial pressure was recorded at baseline conditions and data used for cardiovascular variability and baroreflex sensitivity analysis. Respiratory and behavioral parameters were assessed during this data collection session.ResultsMicroinjections of oxytocin (0.5 µg) into the central nucleus of amygdala produced hypergrooming behavior but did not change cardiorespiratory parameters. However, hypergrooming evoked by microinjections of oxytocin (1 µg) into the central nucleus of amygdala was accompanied by increase in arterial pressure, heart rate and ventilation and augmented the power of low and high (respiratory-related) frequency bands of the systolic arterial pressure spectrum. No changes were observed in power of the low and high frequency bands of the pulse interval spectrum. Baroreflex sensitivity was found lower after oxytocin microinjections, demonstrating that the oxytocin-induced pressor response may involve an inhibition of baroreflex pathways and a consequent facilitation of sympathetic outflow to the cardiovascular system.ConclusionsThe microinjection of oxytocin (1 µg) into the central nucleus of amygdala not only induces hypergrooming but also changes cardiorespiratory parameters. Moreover, specific oxytocin receptor antagonism attenuated hypergrooming but did not affect pressor, tachycardic and ventilatory responses to oxytocin, suggesting the involvement of distinct neural pathways.

18 19 Previously we have demonstrated that microinjection of oxytocin (OT) into the central nucleus of 20 amygdala (CeA) induces hypergrooming in Wistar rats, a model of compulsion. The Wistar 21 Audiogenic Rat (WAR) strain is a genetic model of generalized tonic-clonic seizures. Here we 22 quantified grooming behavior in WAR, with grooming scores, flowcharts and directed graphs of 23 syntactic and non-syntactic grooming chains, after bilateral administration of OT or saline (SAL) into 24 the CeA and investigated the association between hypergrooming and imunohistochemistry of Fos 25 activated compulsion networks and proposing a computational model of grooming behavior. The 26 activated networks, driven from a CeA OT-dependent grooming pattern, in both Wistar and WAR were 27 detected as Fos+ regions: orbitofrontal cortex, striatum, paraventricular nucleus of the hypothalamus, 28 dentate gyrus, substantia nigra compacta and reticulata. In conclusion we can drive hypergrooming in 29 WARs, defined previously as a model of ritualistic motor behavior in Wistar rats, with OT from CeA, a 30 limbic structure and one of the principal amygdala complex outputs. Furthermore, our current pioneer 31 behavioral and cellular description considers that hypergrooming (compulsion) in WARs is a 32 comorbidity because: (1) WARs have the highest grooming scores , when exposed only to novelty (2) 33 WARs have better grooming scores than Wistars after CeA-SAL, (3) WARs perform much better than 34 Wistars in OT-CeA-dependent highly stereotyped behavioral sequences, detected by flowcharts as a 35 combination of syntactic/non-syntactic grooming chains, (4) the behavioral sequences here 36 demonstrated for grooming and hypergrooming can be modeled as quite reliable Markov chains and (5) 37 with the exception of CeA-SAL injected animals, an exquisite map of brain Fos expression was 38 detected in typical cortico-striatal-thalamic-basal ganglia-cortical circuit, among new areas, driven by 39 OT-CeA. 40 41 -900 -Ribeirão Preto -SP 45 -Brazil 46 47 48 Author Summary 49 50 Grooming is a complex set of regular behavioral sequences in rodents that can be mimicked with 51 several pharmacological or molecular biology interventions. We have demonstrated previously that 52 microinjection of the brain peptide oxytocin into the amygdala, a limbic region, induces 53 hypergrooming in Wistar rats, a model of compulsion. The Wistar Audiogenic Rat strain is a genetic 54 model of generalized seizures, in fact a model of epilepsy. Here we quantified grooming behavior in 55 Wistar Audiogenic Rats, using several behavioral tools such as grooming scores, behavioral sequences 56 and graphs of grooming chains, after bilateral administration of the oxytocin or its control (saline) into 57 the amygdala. We also investigated the association between hypergrooming and activation of 58 compulsion networks, proposing a computational (virtual) model of grooming behavior. Basically we 59 were able to detect activated networks, driven from amygdala and the consequent oxytoci...