Annamaria Capozzo

The pedunculopontine nucleus projection to the parafascicular nucleus of the thalamus: an electrophysiological investigation in the rat.

Summary. Extracellular electrophysiological recordings of neurons of the parafascicular nucleus of the thalamus were done in normal rats and in rats bearing lesions of either the cerebellar nuclei or the entopeduncular nucleus to investigate the functional control of the pedunculopontine nucleus on the parafascicular nucleus. A total of 97 neurons were recorded in the parafascicular nucleus in intact rats, 83 in rats bearing a chronic electrolytic lesion of the ipsilateral deep cerebellar nuclei, and 69 in rats bearing an ibotenate lesion of the ipsilateral entopeduncular nucleus. Lesions of the cerebellar nuclei or the entopeduncular nucleus were made to evaluate the participation of cerebellothalamic fibers or of polysynaptic basal ganglia circuits in the responses recorded in parafascicular neurons following electrical microstimulation of the ipsilateral pedunculopontine nucleus. Two types of excitation and one type of inhibition were the main responses observed in neurons of the parafascicular nucleus following stimulation of the pedunculopontine nucleus. The first type of excitation, observed in 49.5% of neurons recorded in normal rats, had an onset of 1.8 ± 0.6 ms, lasted 9.2 ± 0.8 ms and was able to follow high frequency stimulation over 300 Hz. The second type of excitation, observed in a smaller percentage of neurons recorded (3.1%), was a long-latency (8.3 ± 0.7 ms) activation lasting 19.0 ± 4.5 ms. It did not follow stimulation frequencies higher than 50–100 Hz. The inhibitory response was observed in 17.5% of the neurons recorded. The latency of this inhibition was 4.5 ± 1.8 ms and the duration 41.9 ± 6.8 ms. In rats bearing a lesion of the deep cerebellar nuclei or of the entopeduncular nucleus, the short-latency activation was still present in 24.1% and 31.9% of neurons recorded, respectively. However, the occurrence of the long-latency excitation rats bearing lesions of either the cerebellum or the entopeduncular nucleus increased to 12.1% and to 17.4%, respectively, while the occurrence of the inhibition rose to 22.9% and to 28.9%. These results show that an excitatory influence on the parafascicular nucleus is exerted by the pedunculopontine nucleus irrespectively of the presence of cerebellofugal fibers. This influence appears to be also independent from the integrity of basal ganglia circuits having a relay at the level of the entopeduncular nucleus. However, the variety of responses recorded suggests that the influences of the pedunculopontine nucleus on the parafascicular nucleus are by far more complex than those exerted on its basal ganglia targets such as the substantia nigra. The results are discussed according to a model of functioning of pedunculopontine fibers directed to thalamic and basal ganglia nuclei.

The function of the pedunculopontine nucleus in the preparation and execution of an externally-cued bar pressing task in the rat

In the present study the role of the pedunculopontine nucleus (PPN) in the preparation and execution of an externally-cued rewarded motor act was investigated. Animals were instructed to press down a lever at the presentation of a combined visual and acoustic signal and were required to hold down the lever until a trigger stimulus occurred after an unpredictable delay ranging from 2 to 4 s. The trigger stimulus required animals to release the lever and to press a second lever for food reinforcement. The time between instruction and trigger signals represented the preparation phase preceding movement. Unilateral ibotenic acid-induced focal degeneration of pedunculopontine neurons did not influence either reaction and movement times, or capacity of the animals to correctly respond to presentation of stimuli of behavioral significance. On the contrary, bilateral lesions increased both reaction and movement times, and dramatically reduced the percentage of correct responses. The analysis of incorrect responses suggested that the most striking deficit exhibited by the animals following the bilateral lesion was a lack of conditioned response to the signal initiating each trial. However, the animals retained the capability to respond correctly in some trials, and were able to collect the reward when delivered outside the behavioral context. Histological analysis of lesions showed that in addition to loss of neurons within the pedunculopontine region, reduction of tyrosine-hydroxylase positive neurons had occurred in the pars compacta of the substantia nigra. The data suggest that the PPN is involved in the preparation and execution of externally-cued movements, and demonstrate that its destruction mimics the main effects produced by the dopaminergic denervation of the dorsal striatum.

The pedunculopontine tegmental nucleus: implications for a role in modulating spinal cord motoneuron excitability.

There is evidence that deep brain stimulation (DBS) of the pedunculopontine tegmental nucleus (PPTg) improves parkinsonian motor signs. The mechanisms that mediate these effects and the modifications that occur in the PPTg in Parkinson’s disease (PD) are not fully known and are the object of current debate. The aim of this paper was to critically review available data with respect to (1) the presence of PPTg neurons linked to reticulospinal projections, (2) the involvement of these neurons in modulating spinal reflexes, and (3) the participation of fibers close to or within the PPTg region in such modulation. The PPTg neurons are distributed in a large pontotegmental region, stimulation of which can evoke activity in hindlimb, shoulder and neck muscles, and potentiate motor responses evoked by stimulation of dorsal roots. This influence seems to be carried out by fast-conducting descending fibers, which likely run in the medial reticulospinal pathway. It is yet unclear which neurotransmitters are involved and on which elements of the gray matter of the spinal cord PPTg fibers synapse. The modulation of spinal cord activity which can be achieved by stimulating the PPTg region seems to be mediated not only by PPTg neurons, but also by tecto-reticular fibers which run in the pontotegmental area, and which likely are activated during PPTg-DBS. The importance of these fibers is discussed taking into account the degeneration of PPTg neurons in PD and the benefits in gait and postural control that PPTg-DBS exerts in PD. The potential usefulness of PPTg-DBS in other neurodegenerative disorders characterized by neuronal loss in the brainstem is also considered.

Unilateral lesions of the pedunculopontine nucleus do not alleviate subthalamic nucleus-mediated anticipatory responding in a delayed sensorimotor task in the rat.

Lesions of the subthalamic nucleus (STN) in the rat are known to cause anticipated movements in behavioral tasks requiring a preparatory period before the execution of externally cued conditioned movements. In the present study, we describe the effects of lesions of the pedunculopontine nucleus (PPN), a structure located on the outflow of the STN to lower brainstem and spinal motor nuclei, on the anticipatory responding caused by a unilateral lesion of the STN in a delayed sensorimotor task. Rats were instructed to keep a lever pressed down by the presentation of a composite visual and acoustic signal, and were required to hold the lever pressed until a trigger stimulus occurred after an unpredictable delay. The trigger stimulus required the animals to release the lever and to press a second lever for food reinforcement. The task was evaluated according to analysis of movement parameters and errors made by the animals during the preparative and executive phases of the conditioned movement. An ibotenate lesion was placed into the STN in either side of the brain. This lesion was followed 3 weeks later by an ibotenate lesion of the PPN ipsilaterally to the STN previously lesioned. The results indicate that the anticipatory responding induced by the STN lesion was not alleviated by the subsequent PPN lesion. However, the animals bearing the combined lesion were severely impaired in conditioned responding to salient stimuli involved in the paradigm and showed side-specific lengthening of reaction and movement times without global motor impairments. The results suggest that the anticipatory responses caused by STN lesions do not require the intervention of the PPN and that the disruption of the dopaminergic nigrostriatal pathway following the combined lesion may be responsible for impairments observed.

Dopamine denervation of specific striatal subregions differentially affects preparation and execution of a delayed response task in the rat

In the present study, the effects of unilateral or bilateral dopamine denervation of either the dorsal or ventral striatum on the preparation and execution of a delayed response task in the rat were investigated. Animals were instructed to hold a lever pressed down by the presentation of a visual and/or acoustic signal, and were required to hold the lever until a trigger stimulus occurred after an unpredictable delay ranging from 2 to 4 s. The trigger stimulus required animals to release the lever and to press a second lever for food reinforcement. The time between instruction and trigger signal represented the preparation phase preceding movement. The motor performance was evaluated by using reaction and movement times in addition to correct responses in each session. Dopaminergic denervation of either the dorsal or ventral striatum ipsilaterally to the side in which the second lever to be pressed was located did not significantly change reaction and movement times, although it reduced the percentage of correct trials. A significant increase of both reaction and movement times was recorded only after bilateral denervation of the ventral striatum. The analysis of incorrect responses indicated that dopaminergic innervation of the two striatal subregions had different functions in the correct execution of the behavioral paradigm. In the group of animals with dorsal lesions the most frequent incorrect response was represented by a lack of the conditioned response to the presentation of the instruction stimulus starting the trial. If the animals reacted properly to this signal, the performance thereafter was correct in the majority of trials. Conversely, animals with ventral lesions exhibited a large repertoire of incorrect responses throughout the paradigm, including premature release or delayed press of levers, and omission of the second lever press. Histological verification of brain coronal sections by tyrosine-hydroxylase immunoreactivity showed that the lesions were confined in either the dorsal or ventral striatum, sparing the lateral region. The data support the hypothesis that dopaminergic innervation enables the two striatal regions to differently participate in the preparation and execution of complex delayed sensorimotor tasks. Indeed, the dorsal striatum seems to be involved in the correct utilization of external sensory information for the initiation of conditioned behavior, whereas, the ventral striatum appears to be mainly concerned with the temporal expectation of impending stimuli that trigger reward-reinforced movements.

Cholinergic excitation from the pedunculopontine tegmental nucleus to the dentate nucleus in the rat

In spite of the existence of pedunculopontine tegmental nucleus (PPTg) projections to cerebellar nuclei, their nature and functional role is unknown. These fibers may play a crucial role in postural control and may be involved in the beneficial effects induced by deep-brain stimulation (DBS) of brainstem structures in motor disorders. We investigated the effects of PPTg microstimulation on single-unit activity of dentate, fastigial and interpositus nuclei. The effects of PPTg stimulation were also studied in rats whose PPTg neurons were destroyed by ibotenic acid and subsequently subjected to iontophoretically applied cholinergic antagonists. The main response recorded in cerebellar nuclei was a short-latency (1.5-2 ms) and brief (13-15 ms) orthodromic activation. The dentate nucleus was the most responsive to PPTg stimulation. The destruction of PPTg cells reduced the occurrence of PPTg-evoked activation of dentate neurons, suggesting that the effect was due to stimulation of cell bodies and not due to fibers passing through or close to the PPTg. Application of cholinergic antagonists reduced or eliminated the PPTg-evoked response recorded in the dentate nucleus. The results show that excitation is exerted by the PPTg on the cerebellar nuclei, in particular on the dentate nucleus. Taken together with the reduction of nicotinamide adenine dinucleotide phosphate-diaphorase-positive neurons in lesioned animals, the iontophoretic experiments suggest that the activation of dentate neurons is due to cholinergic fibers. These data help to explain the effects of DBS of the PPTg on axial motor disabilities in neurodegenerative disorders.

Behavioural learning‐induced increase in spontaneous GABAA‐dependent synaptic activity in rat striatal cholinergic interneurons

Cholinergic striatal interneurons play a crucial role in cognitive aspects of context‐dependent motor behaviours. They are considered to correspond to the tonically active neurons (TANs) of the primate striatum, which phasically decrease their discharge at the presentation of reward‐related sensory stimuli. The origin of this response is still poorly understood. Therefore, in the present paper, we have investigated whether synaptic changes establish in cholinergic interneurons from young rats that have learned a rewarded, externally cued sensorimotor task. Corticostriatal slices were prepared from both control and trained rats. No significant change in intrinsic membrane properties and evoked synaptic activity was observed in cholinergic interneurons, nor the responsiveness to exogenously applied dopaminergic and glutamatergic agonists was modified. Conversely, an increased occurrence of spontaneous bicuculline‐sensitive depolarizing postsynaptic potentials (sDPSP) was recorded. The frequency of the GABAA‐mediated sDPSP was increased in comparison to not‐conditioned rats. Overall, these results suggest that after learning a rewarded sensorimotor paradigm an increased GABA influence develops on cholinergic interneurons. The origin of this effect might be searched in collaterals of GABAergic output spiny neurons as well as in GABAergic striatal interneurons impinging onto cholinergic interneurons. This intrastriatal mechanism might be involved in the phasic suppression of discharge of TANs at the presentation of reward‐related sensory stimuli.

Fluorescent light induces neurodegeneration in the rodent nigrostriatal system but near infrared LED light does not

We investigated the effects of continuous artificial light exposure on the mouse substantia nigra (SN). A three month exposure of C57Bl/6J mice to white fluorescent light induced a 30% reduction in dopamine (DA) neurons in SN compared to controls, accompanied by a decrease of DA and its metabolites in the striatum. After six months of exposure, neurodegeneration progressed slightly, but the level of DA returned to the basal level, while the metabolites increased with respect to the control. Three month exposure to near infrared LED light (∼710nm) did not alter DA neurons in SN, nor did it decrease DA and its metabolites in the striatum. Furthermore mesencephalic cell viability, as tested by [3H]DA uptake, did not change. Finally, we observed that 710nm LED light, locally conveyed in the rat SN, could modulate the firing activity of extracellular-recorded DA neurons. These data suggest that light can be detrimental or beneficial to DA neurons in SN, depending on the source and wavelength.

Continuous stimulation of the pedunculopontine tegmental nucleus at 40 Hz affects preparative and executive control in a delayed sensorimotor task and reduces rotational movements induced by apomorphine in the 6-OHDA parkinsonian rat.

The pedunculopontine tegmental nucleus (PPTg) relays basal ganglia signals to the thalamus, lower brainstem and spinal cord. Using the 6-hydroxydopamine (6-OHDA) rat model of parkinsonism, we investigated whether deep brain stimulation (DBS) of the PPTg (40 Hz, 60 μs, 200-400 μA) may influence the preparative and executive phases in a conditioned behavioural task, and the motor asymmetries induced by apomorphine. In the conditioned task, rats had to press two levers according to a fixed delay paradigm. The 6-OHDA lesion was placed in the right medial forebrain bundle, i.e. contralaterally to the preferred forepaw used by rats to press levers in the adopted task. The stimulating electrode was implanted in the right PPTg, i.e. contralateral to left side, which was expected to be most affected. The lesion significantly reduced correct responses from 63.4% to 16.6%. PPTg-DBS effects were episodic; however, when rats successfully performed in the task (18.9%), reaction time (468.8 ± 36.5 ms) was significantly increased (589.9 ± 45.9 ms), but not improved by PPTg-DBS (646.7 ± 33.8 ms). Movement time was significantly increased following the lesion (649.2 ± 42.6 ms vs. 810.9 ± 53.0 ms), but significantly reduced by PPTg-DBS (820.4 ± 39.4 ms) compared to sham PPTg-DBS (979.8 ± 47.6 ms). In a second group of lesioned rats, rotations induced by apomorphine were significantly reduced by PPTg-DBS compared to sham PPTg-DBS (12.2 ± 0.6 vs. 9.5 ± 0.4 mean turns/min). Thus, it appears that specific aspects of motor deficits in 6-OHDA-lesioned rats may be modulated by PPTg-DBS.

Cholinergic input from the pedunculopontine nucleus to the cerebellum: implications for deep brain stimulation in Parkinson's disease

Deep brain stimulation (DBS) is a well established electrophysiological treatment initially applied to treat medication-refractory motor symptoms in Parkinsons disease (PD), and is now being explored for several neurological and psychiatric disorders. The specific physiological mechanisms underlying the effectiveness of DBS are not fully understood, although some hypothesized general mechanisms may be acceptable (Wichmann and DeLong, 2016). Early hypotheses suggested that stimulation of the subthalamic nucleus (STN) in PD produced the same clinical effect as a lesion. In other words, DBS was initially considered to suppress or modulate the abnormal bursting discharge patterns that occur in STN neurons in parkinsonian patients. Several mechanisms have been proposed to explain this effect, invoking what would happen at the site of stimulation and/or in the neuronal circuitry to which the targeted region for stimulation is functionally connected. These mechanisms include depolarization block caused by increase of potassium currents, inactivation of sodium channels, presynaptic depression of excitatory afferents, and stimulation-induced activation of inhibitory afferents. However, evidence in favor of activation of STN neurons was also provided, possibly mediated by stimulation-induced activation of excitatory projections from the motor cortex, or by a direct effect of stimulation directly on STN neurons. Moreover, neuronal activity may be phase locked to the pulse train, with following frequencies dictated by the stimulus interval.