In Wistar rats susceptible to audiogenic seizures (Wistar AS) inbred in our laboratory, the exposure to an intense sound induces an epileptic seizure characterized by a running episode followed by a tonic phase showing the major involvement of brainstem structures. After 10-20 sound-induced seizures, development of facial and forelimb clonus and/or tonic-clonic seizures characterize the generalization from brainstem to the forebrain as a result of seizure repetition. In order to specify the anatomical substrates of repeated audiogenic seizures in Wistar AS, we used the 2-deoxyglucose (2DG) technique over a 5 min period to map the midbrain and forebrain structures activated by audiogenic seizures before and after seizure repetition. In naive Wistar AS, six of the 22 structures showed a significant 20-56% increase in relative optical densities compared to non-epileptic controls; these were central and medial amygdala nuclei, perirhinal cortex, medial septum, subthalamic and caudate nuclei. In kindled Wistar AS, 12 additional structures showed a significant 16-121% increase in 2DG labeling. These structures were the substantia nigra, all layers of the hippocampus, the basolateral amygdala, three thalamic nuclei, the frontal motor and prefrontal cortices. In conclusion, the metabolic activation of midbrain and forebrain areas in kindled versus naive Wistar AS rats reflects the changes in the nature of the seizures and the involvement of these structures in the spread of seizure activity from the brainstem to the forebrain during seizure repetition.

Audiogenic seizures, a model of brainstem epilepsy, are characterized by a tonic phase (sustained muscular contraction fixing the limbs in a flexed or extended position) associated with a short cortical electroencephalogram flattening. When sound-susceptible rats are exposed to repeated acoustic stimulations, kindled audiogenic seizures, characterized by a clonic phase (facial and forelimb repetitive jerks) associated with cortical spike-waves, progressively appear, suggesting that repetition of brainstem seizures causes a propagation of the epileptic discharge toward the forebrain. In order to determine the structures through which this propagation occurs, four kinds of experiments were performed in non-epileptic rats and in sound-susceptible rats exposed to single or repeated sound stimulations. The following results were obtained:(1) Electrical amygdalar kindling was similar in non-epileptic and naı̈ve-susceptible rats, but was facilitated in sound-susceptible rats submitted to 40 acoustic stimulations and presenting kindled audiogenic seizures.(2) Audiogenic seizures induced an increase in [14C]2-deoxyglucose concentration in the amygdala after a single seizure, and in the amygdala, hippocampus and perirhinal and piriform cortices after a kindled audiogenic seizure.(3) A single audiogenic seizure induced the expression of c-Fos protein mainly in the auditory nuclei. A few cells were stained in the amygdala. After 5–10 audiogenic seizures, a clear staining appeared in the amygdala, and perirhinal and piriform cortices. The hippocampus expressed c-Fos later, after 40 audiogenic seizures.(4) Injection of lidocaine into the amygdala did not modify single audiogenic seizures, but suppressed myoclonias and cortical spike-waves of kindled audiogenic seizures. Similar deactivation of the hippocampus failed to modify kindled audiogenic seizures.Taken together, these data indicate a critical role for the amygdala in the spread of audiogenic seizures from brainstem to forebrain.

In the present study, we compared Interictal local cerebral metabolic rates for glucose (LCMRglcs) in a strain of audiogenic rats (Wistar AS) selected in our laboratory to Interictal LCMRglcs in a strain of control non-epileptic (NE) rats. Two groups of Wistar AS were studied, one group exposed to a single audiogenic seizure and one group of kindled rats exposed to 40 daily repetitive seizures. Control NE animals were exposed to a single sound exposure which did not induce any behavioral disturbance. Interictal LCMRglcs were measured by the quantitative autoradiographic [ 14C]2-deoxyglucose technique 5 days after the last sound exposure. LCMRglcs were similar in the three groups of rats in 80% of the structures. Compared to the control NE strain, Interictal metabolic levels were mainly decreased in auditory structures of Wistar AS, either naive or kindled, thus confirming auditory impairment in audiogenic animals. LCMRglcs were increased over control levels in both groups of Wistar AS in cerebellar regions. This increase of cerebellar functional activity in Wistar AS compared to control NE rats might reflect an increased cerebellar input which, together with auditory impairment, may facilitate the induction of seizure activity in Wistar AS. Finally, there was no difference between the Interictal cerebral metabolic level of naive and kindled Wistar AS, except in the cerebellar dentate nucleus where LCMRglc was significantly higher in kindled than in naive animals.

The quantitative autoradiographic [14C]-iodoantipyrine technique was applied to measure the effects of a 30-min period of pentylenetetrazol (PTZ)-induced status epilepticus (SE) on local cerebral blood flow (LCBF) in rats 10 (P10), 14 (P14), 17 (P17), and 21 (P21) days after birth. The animals received repetitive, timed injections of subconvulsive doses of PTZ until SE was reached. At P10, SE induced a 32 to 184% increase in the rates of LCBF affecting all structures studied. In P14- and P17 PTZ-treated rats, LCBF values significantly increased in two-thirds of the structures belonging to all systems studied and were not changed by SE in the parietal cortex, dorsal hippocampus, and dentate gyrus. At P21, rates of LCBF were still increased in 48 of the 73 structures studied; however, LCBF values were decreased by SE in most cortical areas, the hippocampus, and the dentate gyrus. CBF and cerebral metabolic rate for glucose (CMRglc) remained coupled in both controls and PTZ-exposed rats. Our results show that changes in LCBF with seizures are age dependent. At the most immature ages, P10 and P14, both LCBF and local CMRglc (LCMRglc) values are largely increased by long-lasting seizures. At P17 and P21, the blood flow response to SE becomes more heterogeneous, with specific decreases in the hippocampus and cortex at P21. The absence of mismatch between LCBF and LCMRglc in PTZ-exposed rats at all ages may explain at least partly why the immature brain is more resistant to seizure-induced brain damage than the adult brain.

An experimental model of status epilepticus has been developed in the immature rat by administration of pentylenetetrazol (PTZ) using repetitive, timed intraperitoneal injections of subconvulsive doses. The pattern of behavioral signs has been well characterized in each age group, i.e. 10 (P10), 14 (P14), 17 (P17) and 21 postnatal days (P21). In this model, the dose of convulsant could be adjusted as a function of interindividual sensitivity and status epilepticus lasted for quite a long duration to allow the measurement of local cerebral metabolic rates for glucose (LCMRglc) by means of the [14C]2-deoxyglucose method [J. Neurochem., 28 (1977) 897–916]. To estimate LCMRglc during status epilepticus, the lumped constant (LC) was re-calculated in controls and PTZ-treated rats. The control LC was 0.54 at P10 and 0.50–0.51 at the three older ages studied (P14, P17 and P21). During status epilepticus, it increased to 0.64 in P10 rats and decreased to 0.42 and 0.40, respectively, in P17 and P21 animals. At P14, LC was not affected by seizures. The measurements of brain lactate levels showed a large 4.5–10-fold increase in PTZ-treated rats as compared to controls at all ages. The results of the present study show that the immature brain responds to sustained seizure activity in a specific way according to its postnatal age. Moreover, our results underscore the necessity of re-calculation of LC to the quantification of LCMRglc in such pathological states, particularly in immature animals.

The quantitative autoradiographic [14C]2-deoxyglucose technique (2DG) was applied to measure the effects of pentylenetetrazol (PTZ)-induced status epilepticus (SE) on local cerebral metabolic rates for glucose (LCMRglc) in 10 (P10)-, 14 (P14)-, 17 (P17)- and 21 (P21)-day-old rats. To produce long-lasting SE (55 min), the animals received repetitive, timed intraperitoneal injections of subconvulsive doses of PTZ until SE was reached. At P10 and P14, SE induced a marked increase in LCMRglc which affected 66 of the 76 structures studied. Increases were especially high (200–400%) in limbic and motor cortices at P10 and in some brainstem areas at these 2 ages. At P17 and P21, average brain glucose utilization was similar in seizing and control rats, but in PTZ-treated rats reflected a redistribution in local metabolic rates with increases in brainstem, midbrain, hypothalamus and septum, decreases in cortex, hippocampus, some sensory areas and white matter and no change in many motor and limbic structures. In a few cerebral regions, such as hippocampus, dentate gyrus and mammillary body, LCMRglc did not increase at P10 and P14 and decreased at P17 and P21 in PTZ- vs. saline-treated rats. The results of the present study show that the immature brain responds to sustained seizure activity in a specific way according to its maturational state. Moreover, these data allow the mapping of the vulnerability of cerebral structures to seizures, according to their metabolic response to convulsions.

Adenosine is now considered as a major regulatory agent in the mammalian central nervous system. Its actions are mediated by specific receptors which are coupled with an adenylate cyclase system via a G protein. The postnatal development of adenosine A1 receptors was studied by quantitative autoradiography using [H-3]N6-cyclohexyladenosine, a potent receptor agonist in 42 rat brain structures. The coupling of these sites to G proteins was examined by measuring the effects of in vitro addition of guanylyl-5'-imidodiphosphate, a stable analogue of guanosine triphosphate, on N6-cyclohexyladenosine binding. [H-3]N6-Cyclohexyladenosine-specific binding was quite low at birth, around 10% of adult levels, and exhibited a rather homogeneous distribution pattern, except in thalamic nuclei. Data showed a sequential development of adenosine A1 receptors in relation to the course of maturation of cerebral structures with a proliferation peak which paralleled rapid brain growth.The time period by which adult levels are reached differed according to the cerebral region studied. N6-Cyclohexyladenosine-specific binding sites appeared to be functionally linked to G proteins in all structures and at all postnatal stages. However, the potency of guanylyl-5'-imidodiphosphate to displace N6-Cyclohexyladenosine binding was significantly lower before 5 days of age, suggesting functional changes during postnatal maturation in cerebral pathways modulated by adenosine.

An autoradiographic method has been developed for the regional assessment of cerebral tracer levels after the acute intravenous injection of [3-C-14]beta-hydroxybutyrate in developing rats. The animals were studied at five postnatal stages, i.e. postnatal day 10 (P10), P14, P17, P21 and P35. Tracer levels were high from P10 to P17, reaching peak values at P14, which were two- to threefold higher than those at P10. At P17, tracer concentrations were about twice as low as at P14. Between P17 and P21, regional C-14 concentrations were again reduced by about twofold in all areas studied and decreased further by about 50% after weaning reaching quite low levels by P35. The distribution of C-14 inside sections appeared to be rather homogeneous throughout the brain at all stages studied, never exceeding a ratio higher than 2 at any stage studied.These results are in good agreement with previous data on the rate of uptake and utilization of beta-hydroxybutyrate by the immature rat brain.

The influence of an early chronic phenobarbital (PhB) exposure on local cerebral glucose utilization (LCGU) and on behavior was studied in the rat. The animals were treated from Postnatal Day 2 to Postnatal Day 35 by a daily injection of 50 mg/kg PhB or by saline and tested between 10 and 35 days for short-term effects of the drug on LCGU and between 70 and 90 days for long-term effects of PhB on LCGU and behavior. PhB induced short- and long-lasting reductions in the overall rates of LCGU in hippocampal and cerebellar areas, but no significant changes in LCGU in the different cell layers of these two cerebral areas. PhB also changed the pattern of maturation of the rates of LCGU as compared to control subjects. The barbiturate treatment induced a decrease in the exploratory behavior of PhB- as compared to saline-treated rats in the open field, as well as a significant 25% decrease in the rate of spontaneous alternation with delay. In addition, PhB-treated rats needed significantly more time than control animals to perform their trials in the nonrewarded T maze testing. However, the neonatal barbiturate exposure did not induce changes in performances of adult rats in the rewarded eight arm maze. The results of the present study show that there is no apparent correlation between the rates of energy metabolism in the hippocampus and the impairment of learning abilities of adult rats in behavioral tests related to the hippocampus.

The influence of a chronic phenobarbital (PhB) treatment on postnatal evolution of local cerebral metabolic rates for glucose (LCMRglc) was studied in 58 cerebral structures of freely moving rats. The animals received a daily subcutaneous injection of PhB at a dose of 50 mg/kg between days 2 and 35 or an equivalent volume of saline for controls and were studied at 5 postnatal stages, i.e. 10, 14, 17, 21 and 35 days, and at the adult stage. Body and brain weights were both reduced by 6-21% over the whole period studied. PhB exposure induced significant decreases in LCMRglc during the period of pharmacological treatment, i.e. until 35 days, except at the stage of 17 days as well as long-term reductions in LCMRglc of adult rats in 36 out of the 58 brain regions studied. These decreases affected all systems studied, sensory systems as well as limbic, hypothalamic, motor and white matter areas. In addition to a growth retardation, PhB also seemed to be able to induce a delay in the acquisition of auditory function which matures early during postnatal life. The long-term deficits in cerebral energy metabolism due to PhB in the adult rat also confirm the behavioral deficits which have been shown previously after early PhB exposure.

The postnatal changes in local cerebral blood flow in freely moving rats were measured by means of the quantitative autoradiographic [14C]iodoantipyrine method. The animals were studied at 10, 14, 17, 21 and 35 days and at the adult stage. At 10 days after birth, rates of blood flow were very low and quite homogeneous in most cerebral structures except in a few posterior areas. From these relatively uniform levels, values of local cerebral blood flow rose notably to reach a peak at 17 days in all brain regions studied. Rates of blood flow decreased between 17 and 21 days after birth and then increased from weaning time to reach the known characteristic distribution of the adult rat. The postnatal evolution of local cerebral blood in the rat is in good agreement with previous studies in other species such as dog and humans that also show higher rates of cerebral blood flow and glucose utilization at immature stages. However, in the rat, local cerebral blood flow and local cerebral glucose utilization are not coupled over the whole postnatal period studied, since blood flow rates reach peak values at 17 days whereas glucose utilization remains still quite low at that stage. The high rate of cerebral blood flow in the 17-day-old rat may reflect the energetic and biosynthetic needs of the actively developing brain that are completed by the summation of glucose and ketone body utilization.