Dan C. McIntyre

Transfer, interference and spontaneous recovery of convulsions kindled from the rat amygdala☆

Abstract Bipolar electrodes were implanted into the amygdala of each hemisphere of adult male rats. A short burst of low-intensity stimulation was applied to one of these electrodes once each day. Initially there was little response. With repetition, epileptiform responses progressively developed until each daily stimulus triggered a behavioral convulsion (kindling effect). Following six convulsions, the procedure was applied to the contralateral hemisphere. Convulsions were observed to kindle more rapidly, especially if a rest interval of 2 weeks followed the last primary site convulsion. When stimulation was reapplied to the primary site, convulsions were not triggered. This was associated with failure to evoke local after-discharge and/or failure of the after-discharge to propagate. Several trials were necessary to re-establish convulsion, unless a 2 week rest preceded testing, in which case convulsions were triggered on the first trial. Following a series of convulsions triggered from either hemisphere, the contralaterally triggered convulsions, when they appeared, showed consistently longer onset lattencies. These latency shifts were attentuated when rest intervals preceded the testing. Latency shifts and seizures failures were not observed if the preceeding series of convulsions was reduced from six to one. Lesions at the tip of either electrode had little effect on the results obtained in the contralateral hemisphere. Together, the results imply that: kindling establishes a lasting trace which is both transynaptic and widespread, kindling from a second location establishes a second trace utilizing parts of the existing trace, a series of convulsions leaves a less durable after-effect with a decay time of about 2 weeks and which interferes with various aspects of seizure activity, and the trace which activates the convulsions is less susceptible to interference from the after-effect.

Kindling mechanisms: current progress on an experimental epilepsy model.

For many years a major goal of epileptologists has been to discover the mechanisms of seizure generation. The experimental approaches to the problem have been numerous, addressing acute and chronic determinants in both focal and non-focal models. Perhaps the best controlled model of chronic focal epilepsy is that of kindling (Goddard et al., 1969). The latter refers to the progressive development of electroencephalographic and behavioral seizure resulting from the repeated application of a low-intensity electrical stimulus to one of many discrete forebrain structures. The increase in manifestations of the motor seizure proceeds through several stages (Racine, 1972). With subcortical stimulation, there is an initial ictus of the ipsilateral facial and neck muscles (stages 1 and 2) which is associated with focal EEG seizure. Further stimulations result in secondary generalization of the seizure to involve the forelimbs and trunk (stages 3 and 4), with an additional loss of balance during strong ictus (stage 5). The neurological changes which underlie these behavioral alterations seem to be permanent since animals which have not been stimulated for many months after stage 5 kindling often respond with a full seizure immediately upon reexposure to the original kindling stimulus (Goddard et al., 1969; Wada and Sato, 1974). In this paper, we review selectively some of the more obvious hypotheses concerning the nature of the mechanisms underlying kindling and conclude with a brief summary of the current working hypothesis. For more general or extensive reviews of the kindling literature see Racine (1978), McNamara et al. (1980), Kalichman (1982), and Peterson and Albertson (1982).

Kindling modulates the IL-1β system, TNF-α, TGF-β1, and neuropeptide mRNAs in specific brain regions

Cytokines and neuropeptides may be involved in seizure-associated processes. Following amygdala kindling in rats, we determined alterations of IL-1β, IL-1 receptor antagonist (IL-1Ra), IL-1 receptor type I (IL-1RI), IL-1 receptor accessory proteins (IL-1R AcPs) I and II, TNF-α, TGF-β1, neuropeptide Y (NPY), glycoprotein 130 (gp 130) and pro-opiomelanocortin (POMC) mRNA levels in the parietal, prefrontal and piriform cortices, amygdala, hippocampus and hypothalamus. Messenger RNAs expression in all brain regions was determined 2 h or 3 weeks following the last generalized convulsive seizure triggered from the ipsilateral kindled amygdala. The same brain region sample was used to assay for changes of all mRNA components. The results show that the 2 h-kindled group exhibited a significant up-regulation of IL-1β, IL-1RI, TNF-α and TGF-β1 mRNAs in all three cortical brain regions, amygdala and hippocampus. The largest up-regulation occurred in the prefrontal cortex (about 30-fold induction for IL-1β and TNF-α mRNAs). IL-1R AcP I and II mRNA levels were also up-regulated in the cortical regions. No changes in IL-1β, IL-1RI or TNF-α mRNA levels occurred in the 3 week-kindled group. NPY mRNA levels increased in the hippocampus, prefrontal and piriform cortices in the 2 h-kindled group, while IL-1Ra, gp 130, or POMC mRNA levels did not change in any group. The overall profile of mRNA changes shows specificity of transcriptional modulation induced by amygdala kindling. The data support a role of cytokines and NPY in the adaptive mechanisms associated with generalized seizure activity, with implications for neuroprotection, neuronal dysfunction and vulnerability associated with epileptic activity.

A new model of partial status epilepticus based on kindling

In this experiment, a new model of partial status epilepticus (SE) is described which is based on the antecedent development of a kindled focus. Following kindling, the amygdala was stimulated continuously for 60 min with the previous kindling stimulus (60 Hz sine wave, 50 microA peak-to-peak). This treatment provoked SE in 22 of 35 rats. Without drug intervention, rats spontaneously recovered (SR group) from the seizure between 10 and 24 h. After recovery from SE, after discharge (AD) thresholds were elevated and remained so for the 2 weeks before sacrifice. The histologies of these SR rats indicated massive gliosis and degeneration of the ipsilateral hemisphere, extending from the medial olfactory bulb, through the amygdala-pyriform cortex to the ventral hippocampus. Damage was observed frequently in the midline thalamic nuclei and hippocampal CA1 fields. Interruption of the SE with Nembutal 30 min after the stimulation offset (30 Min group) was occasionally associated with slight gliosis at the kindled electrode, whereas interruption after 4 h of SE (4 Hr group) resulted in more extensive cell loss. The AD thresholds of the 30 Min group, like those of the rats which did not develop SE (NSE group), returned to near-normal values by 2 weeks after SE; only the NSE rats exhibited generalized seizures to their AD threshold stimulus. This model of SE results in brain pathology similar to that found in other models, but has the advantage of being uncontaminated by exogenous chemicals and toxins.

Efferent projections of the anterior perirhinal cortex in the rat.

Because convulsive seizures develop very rapidly from kindling sites in the anterior perirhinal cortex, we studied perirhinal efferents by using the anterograde tracer Phaseolus vulgaris leucoagglutinin (PhAL). PhAL injections into the anterior perirhinal cortex labelled a prominent network of fibers within the frontal cortex that was most dense within layers I and II and layer VI. As individual PhAL injection sites within the perirhinal cortex were restricted to one or two adjacent laminae, we were able to determine that layer V was the main source of the perirhinofrontal projection. This was confirmed by frontal cortex injections of the retrograde tracer Fluorogold (FG).

Potentiation of amygdala kindling in adult or infant rats by injections of 6-hydroxydopamine

Abstract Rats with bilateral amygdala electrodes were pretreated with 6-hydroxydopamine and/or desmethylimipramine (DMI) to deplete norepinephrine (NE) and/or dopamine (DA). Subsequently they were subjected to kindling procedures in which daily low-intensity amygdala stimulation resulted eventually in the development of clonic motor seizures. The NE depletion, but not the DA depletion, resulted in a marked reduction in the number of stimulations required to kindle convulsions (i.e., faster kindling), but increased the latency to onset of convulsions; the convulsion or after-discharge durations were unaffected. Interestingly, the control animals that received DMI alone manifested no obvious permanent neurochemical effect yet exhibited retarded after-discharge evolution. The above data were replicated in two experiments using adult- and infant-injected rats. These data confirm previous reports that the central catecholamines are capable of inhibiting a variety of forms of seizure.

Development of kindling-prone and kindling-resistant rats: selective breeding and electrophysiological studies

Because of the growing need for an animal model of complex partial seizures based on a genetic predisposition, we combined the kindling model of epilepsy with selective-breeding procedures to develop two new lines (or strains) of rats that are kindling-prone or kindling-resistant. The selection of these strains was based on their rates of amygdala kindling. From a parent population of Long Evans hooded and Wistar rats, the males and females that showed the fastest and slowest amygdala kindling rates were selected and bred. Similar selection procedures continued through F11, although there was little or no overlap in the distribution of kindling rates for the two new strains (FAST and SLOW) by F6. Examination of both local and propagating seizure profiles of the new strains from F6 to F10 revealed that the FAST and SLOW rats had similar amygdala afterdischarge (AD) thresholds and associated AD durations. Also, the convulsion profiles of the stage-5 responses were similar, although the severity was greater in the FAST rats. Clearly the selection was not based on local mechanisms controlling the threshold for amygdala AD evocation, but rather for the spread of AD from the focus and the recruitment of other structures, ultimately triggering convulsive seizures. Although evoked potentials and potentiation effects were similar between the strains, the SLOW rats showed a greater paired-pulse depression, raising the possibility that they differ in inhibitory mechanisms. The specificity of strain differences for the amygdala and its associated networks is described in our accompanying paper (McIntyre et al., 1999. FAST and SLOW amygdala kindling rat strains: Comparison of amygdala, hippocampal, piriform and perirhinal cortex kindling. Epilepsy Res. 35, 197-209). These strains should provide many clues to the dispositional differences between individuals for the development of epilepsy originating in temporal lobe structures.

Animal Models of Limbic Epilepsies: What Can They Tell Us?

In this review, we have provided an overview of the implementation and characteristics of some of the most prevalent models of temporal lobe epilepsy in use in laboratories around the world today. These include spontaneously seizing models with status epilepticus as the initial precipitating injury (including the kainate, pilocarpine, and electrical stimulation models), kindling, and models of drug refractoriness. These models share various features with one another, and also differ in many aspects, providing a broader representation of the full spectrum of clinical limbic epilepsies. We have also provided a brief introduction into how animal models of temporal lobe epilepsy facilitate use of modern state-of-the-art techniques in neurobiology to address critical questions in the pathogenesis of epilepsy.

FAST and SLOW amygdala kindling rat strains: comparison of amygdala, hippocampal, piriform and perirhinal cortex kindling

In our companion paper, we selectively bred offspring of a Long Evans Hooded and Wistar rat cross for either fast or slow rates of amygdala kindling (Racine et al., 1999. Development of kindling-prone and kindling resistant rats: Selective breeding and electrophysiological studies, Epilepsy Res. 35, 183-195). Within 10 generations, there was no overlap in the distribution of kindling rates between these newly developed FAST and SLOW kindling strains. In the present report, we compared the local excitability, kindling rates, and convulsion profiles of kindling sites in either the amygdala, dorsal hippocampus, piriform cortex or perirhinal cortex in the two strains. Local excitability, measured as the local afterdischarge (AD) threshold and its duration, showed varied effects between structures and strains. Before kindling, the AD threshold was lower in the FAST than the SLOW rats in the hippocampus, piriform and perirhinal cortices, but not the amygdala (the selection structure). Also, the duration of the AD threshold duration was significantly longer in the FAST than in the SLOW rats in all structures, except the CA1 hippocampus. Most of these differences were maintained after kindling. Kindling itself was significantly faster in the FAST compared with the SLOW rats in all structures; however, the different structural kindling rates showed proportional differences between strains that were about five times different in the amygdala compared with only about two times different in the hippocampus. This suggested a selection bias for the amygdala and its networks. As in other rat strains, the fastest kindling rates were seen in the perirhinal cortex followed by the piriform cortex, amygdala and hippocampus in both FAST and SLOW rats. Other important differences between strains and structures occurred in the stage-5 convulsion profiles, including latency to forelimb clonus, clonus duration and duration of associated local afterdischarges. The differences in kindling profiles between strains and structures were discussed with respect to possible underlying mechanisms, significance for epileptogenesis, and impact on other normal behaviours.

Another inexpensive headplug for the electrical recording and or stimulation of rats

Abstract This paper describes a durable, inexpensive micro-miniature headplug which has been used successfully for centrally stimulating and recording from rats performing vigorous movements.