Karen L. Hoskin

Expression of c-Fos-like immunoreactivity in the caudal medulla and upper cervical spinal cord following stimulation of the superior sagittal sinus in the cat

Migraine is an episodic vascular headache with a well-recognized clinical picture but a poorly understood pathogenesis. Stimulation of a pain-sensitive trigeminally innervated intracranial structure, the superior sagittal sinus (SSS), was undertaken to map the higher-order neurons potentially involved in the processing of vascular head pain. The animals were prepared for stimulation by exposure of the sinus and then maintained under alpha-chloralose anaesthesia for 24 h before SSS stimulation, perfusion and immunohistochemical processing for the detection of Fos protein. Examination of the medulla and upper cervical cord revealed marked increases in Fos-like immunoreactivity in laminae I and IIo of the trigeminal nucleus caudalis and the dorsal horn of the upper cervical spinal cord. In addition, Fos-like immunoreactivity was observed in lamina X of the upper cervical spinal cord, in the commissural and medial nuclei of the solitary tract and in the nucleus retroambigualis. The use of immunohistochemical detection of Fos has allowed visualization of several populations of neurons likely to be involved in the central neural processing of vascular headache syndromes, particularly migraine.

The distribution of trigeminovascular afferents in the nonhuman primate brain Macaca nemestrina: a c-fos immunocytochemical study

An understanding of migraine must be based on data concerning the anatomy and physiology of the pain‐sensitive intracranial structures. Stimulation of the superior sagittal sinus produces changes in brain blood flow and changes in neuropeptide levels similar to those seen in humans during migraine. To better understand the anatomy of the central ramifications of pain‐sensitive intracranial structures we have examined the distribution of c‐fos immunoreactivity in the monkey when the sinus is stimulated. Six adult Macaca nemestrina monkeys were anaesthetised. The superior sagittal sinus was isolated after a midline craniotomy and a paraffin well created. At 24 h after completion of the surgery the sinus was stimulated electrically for 1 h and the brain subsequently removed and processed for c‐fos. In control animals in which the sinus was isolated but not stimulated there was a small amount of c‐fos expression in the caudal brainstem and upper cervical spinal cord. Stimulation of the superior sagittal sinus evoked expression of c‐fos in the caudal superfical laminae of the trigeminal nucleus and in superficial laminae of the dorsal horn of the C1 level of the upper cervical spinal cord. A lesser amount of c‐fos was seen at C2 while no significant labelling above control was observed at C3. These data, while largely confirming the results from the cat concerning the central distribution trigeminovascular afferents, underscore a possibly unique specialisation of trigeminovascular afferents at the C1 level. Given the close evolutionary relationship of the monkey to man it is likely that the cells described in this study represent for primates the nucleus that mediates the pain of migraine.

Inhibition by sumatriptan of central trigeminal neurones only after blood‐brain barrier disruption

1 The 5‐hydroxytryptamine (5‐HT1)‐like agonist, sumatriptan, is highly efficient in the relief of migraine headache and its accompanying symptoms. 2 Experimental evidence has indicated that its site of action may be on the cranial vessels or on the trigeminal innervation of the cranium, or both, since sumatriptan does not pass the blood‐brain barrier easily under normal circumstances. It is, however, not clear whether the blood‐brain barrier is normal or abnormal during a migraine attack. 3 In this study, single unit activity and trigeminal somatosensory evoked potentials in central trigeminal neurones were monitored during electrical stimulation of the superior sagittal sinus. 4 Intravenous administration of sumatriptan (100 μg kg−1) did not alter trigeminal evoked activity unless the permeability of the blood‐brain barrier had been increased by infusion of an hyperosmolar mannitol solution. After blood‐brain barrier disruption, sumatriptan decreased the peak‐to‐peak amplitude of evoked potentials by 40 ± 6% and the probability of firing of single units by 30 ± 9%. Mannitol infusions alone in control animals caused no changes in evoked potentials or single unit activity. 5 The data suggest that in normal circumstances sumatriptan does not have sufficient access to trigeminal neurones to alter their function.

Inhibition of trigeminal neurons by intravenous administration of the serotonin (5HT)1B/D receptor agonist zolmitriptan (311C90): are brain stem sites therapeutic target in migraine ?

&NA; Migraine is a common and debilitating condition. Its treatment has received considerable attention in recent times with the introduction into clinical use of the serotonin (5HT)1B/D‐like agonist sumatriptan. It is known from human studies that the intracranial blood vessels and dura mater are important pain‐sensitive structures since mechanical or electrical stimulation of these vessels, such as the superior sagittal sinus, causes pain. We have developed a model of craniovascular pain by stimulating the superior sagittal sinus and monitoring trigeminal neuronal activity using electrophysiological techniques. In this study we determined the effect of intravenous administration of the novel anti‐migraine compound zolmitriptan (311C90) upon evoked neuronal activity in trigeminal neurons. Nine adult cats were anaesthetised with &agr;‐chloralose (60 mg/kg, i.p.; 20 mg/kg, i.v., 2‐hourly) with all surgery being conducted under halothane (1–3%). The superior sagittal sinus was isolated for electrical stimulation. Recordings were made from caudal trigeminal neurons at the C2 level of the cervical spinal cord with tungsten‐in‐glass microelectrodes. Signals were amplified and analysed by a custom‐written program that enabled software filtering and extraction of both evoked potential and single cell data. Data were collected before and after administration of zolmitriptan. Electrical stimulation of the superior sagittal sinus resulted in activation of neuronal elements within the trigeminal nucleus that could be monitored as single unit activity or as evoked potentials, the latter reflecting both primary afferent and trigeminal cell body activity. The evoked potential recorded from the trigeminal nucleus was 207 ± 14 &mgr;V and was reduced by zolmitriptan (100 &mgr;g/kg, i.v.) to a mean of 98 ± 17 &mgr;V. Similarly, the probability of firing for trigeminal neurons was reduced from a control level of 0.63 ± 0.1 to 0.13 ± 0.05 after a dose of 100 &mgr;g/kg intravenously. These effects weredose‐dependent and were significantly different from the effect of vehicle (P < 0.05). These data demonstrate that systemically administered zolmitriptan can inhibit evoked trigeminovascular activity within the trigeminal nucleus. This inhibition of trigeminal activity may play a role in the anti‐migraine actions of this compound and offers the prospect of a third pathophysiologically consistent target site for anti‐migraine drug effects.

Nitric oxide synthesis couples cerebral blood flow and metabolism

The most fundamental aspect of the cerebral circulation is the well-described coupling of cerebral metabolic activity and cerebral blood flow. A number of substances have been proposed to link flow and metabolism, including K+, pH and adenosine. In the alpha-chloralose anaesthetised cat we studied simultaneously cerebral neuronal activity and local blood flow to attempt to dissociate the two and thus determine the coupling substance. Neuronal activity was determined by monitoring unit firing in the parietal cortex with tungsten in glass microelectrodes while local cerebral blood flow in the same area was monitored continuously using laser Doppler flowmetry. To initiate an increase in metabolic activity and, pari passu, blood flow spreading depression was elicited by needle stick injury. Spreading depression when initiated causes a wave of depolarization, measured as an increased firing rate and associated marked (400 +/- 95%) increase in local cerebral blood flow. Intravenous administration of NG-nitro-L-arginine methyl ester (1-NAME), a potent nitric oxide synthase inhibitor, produced a complete blockade of the hyperemia associated with spreading depression but no change in either resting cell firing or spreading depression-evoked increases in firing rate. These data demonstrate at least for spreading depression-elicited increases in metabolic activity, that nitric oxide (NO) is a key coupling compound that links changes in cerebral blood flow and metabolism. These data imply that NO may have a more general role in flow/metabolism coupling and further studies in other situations are required to determine the extent to which NO is responsible for this fundamental cerebrovascular phenomenon.

Stimulation of the greater occipital nerve increases metabolic activity in the trigeminal nucleus caudalis and cervical dorsal horn of the cat

&NA; Patients with primary headache syndromes often describe a distribution of pain that involves both frontal and occipital parts of the head. Such a distribution of pain does not respect the cutaneous sensory innervation of the head which would divide it into anterior (trigeminally innervated) and posterior (spinal nerve root innervated) regions. Studies of pain‐producing intracranial structures, such as the superior sagittal sinus, have demonstrated that second order neurons as caudal as C2 are activated after either electrical or mechanical stimulation. For this study cats were anaesthetised with halothane (during surgery) and &agr;‐chloralose (60 mg/kg, i.p., then 20 mg/kg intravenous maintenance), paralysed (gallamine 6 mg/kg) and ventilated. The greater occipital nerve was isolated bilaterally and stimulated unilaterally using hook electrodes with stimuli of 100 V at 0.3 Hz. Metabolic activity in the caudal brain stem and upper cervical cord was measured using 2‐deoxyglucose autoradiography and quantitative densitometry. Stimulation of the greater occipital nerve increased metabolic activity by 220% ipsilateral to stimulation and by a lesser amount contralaterally. Increases in metabolic activity were seen in the dorsal horn at the level of C1 and C2 as might be predicted from the cervical origin of the nerve. Neuronal activation appeared contiguous with the trigeminal nucleus caudalis and was in the same distribution as is seen when trigeminally‐innervated structures are stimulated. These data suggest that the well recognised clinical phenomenon of pain at the front and back of the head and in the upper neck are likely to be a consequence of overlap of processing of nociceptive information at the level of the second order neurons.

STIMULATION OF THE MIDDLE MENINGEAL ARTERY LEADS TO FOS EXPRESSION IN THE TRIGEMINOCERVICAL NUCLEUS : A COMPARATIVE STUDY OF MONKEY AND CAT

The pain of a migraine attack is often described as unilateral, with a throbbing or pulsating quality. The middle meningeal artery (MMA) is the largest artery supplying the dura mater, is paired, and pain‐producing in humans. This artery, or its branches, and other large intracranial extracerebral vessels have been implicated in the pathophysiology of migraine by theories suggesting neurogenic inflammation or cranial vasodilatation, or both, as explanations for the pain of migraine. Having previously studied in detail the distribution of the second order neurons that are involved in the transmission of nociceptive signals from intracranial venous sinuses, we sought to compare the distribution of second order neurons from a pain‐producing intracranial artery in both monkey and cat. By electrically stimulating the middle meningeal artery in these species and using immunohistochemical detection of the proto‐oncogene Fos as a marker of neuronal activation, we have mapped the sites of the central trigeminal neurons which may be involved in transmission of nociception from intracranial extracerebral arteries. Ten cats and 3 monkeys were anaesthetised with α‐chloralose and the middle meningeal artery was isolated following a temporal craniotomy. The animals were maintained under stable anaesthesia for 24 h to allow Fos expression due to the initial surgery to dissipate. Following the rest period, the vessel was carefully lifted onto hook electrodes, and then left alone in control animals (cat n = 3), or stimulated (cat n = 6, monkey n = 3). Stimulation of the left middle meningeal artery evoked Fos expression in the trigeminocervical nucleus, consisting of the dorsal horn of the caudal medulla and upper 2 divisions of the cervical spinal cord, on both the ipsilateral and contralateral sides. Cats had larger amounts of Fos expressed on the ipsilateral than on the contralateral side. Fos expression in the caudal nucleus tractus solitarius and its caudal extension in lamina X of the spinal cord was seen bilaterally in response to middle meningeal artery stimulation. This study demonstrates a comparable anatomical distribution of Fos activation between cat and monkey and, when compared with previous studies, between this arterial structure and the superior sagittal sinus. These data add to the overall picture of the trigeminovascular innervation of the intracranial pain‐producing vessels showing marked anatomical overlap which is consistent with the often poorly localised pain of migraine.

Fos expression in the trigeminocervical complex of the cat after stimulation of the superior sagittal sinus is reduced by l-NAME

Primary neurovascular headaches, such as migraine and cluster headache probably involve activation of trigeminovascular pain structures projecting to the trigeminocervical complex of neurons in the caudal brain stem and upper cervical spinal cord. It has recently been demonstrated that blockade of the synthesis of nitric oxide (NO) by an NO synthesis inhibitor can abort acute migraine attacks and thus it is of interest to determine whether there is an influence of NO generation on trigeminocervical neurons. Cats were anaesthetised with alpha-chloralose (60 mg/kg, i.t.). supplemental 20 mg/kg, intravenously (i.v.)) and halothane for surgery (0.5-3% by inhalation). A circular midline craniotomy was performed to isolate the superior sagittal sinus (SSS) for electrical stimulation (0.3 Hz, 150 V, 250 micros duration for 2 h). Two groups were compared, one stimulated after administration of vehicle and the other stimulated after administration of N(G)-nitro-L-arginine methylester (L-NAME: 100 mg/kg, i.v.). After stimulation of the SSS Fos immunoreactivity was observed in lamina I/IIo of the trigeminal nucleus caudalis and dorsal horns of C1 and C2 to a median total of 136 cells (range 122-146). After L-NAME treatment Fos expression was significantly reduced to 40 cells (24-54; P < 0.02). In conclusion, inhibition of NO synthesis L-NAME markedly reduces Fos expression in the trigeminocervical complex of the cat. These data taken together with the clinical observations of the effect of NO synthesis blockade in migraine suggest a role for NO generation in mediating nociceptive transmission in acute migraine.

Fos expression in the midbrain periaqueductal grey after trigeminovascular stimulation

There is an accumulating body of evidence suggesting that the periaqueductal grey (PAG) is involved in the pathophysiology of migraine. Positron emission tomography (PET) studies in humans have shown that the caudal ventrolateral midbrain, encompassing the ventrolateral PAG, has activations during migraine attacks. The PAG may well be involved not only through the descending modulation of nociceptive afferent information, but also by its ascending projections to the pain processing centres of the thalamus. In this study the intranuclear oncogene protein Fos was used to mark cell activation in the PAG following stimulation of the trigeminally‐innervated superior sagittal sinus (SSS) in both cats and in nonhuman primates (Macaca nemestrina). Fos expression in the PAG increased following stimulation to a median of 242 cells (interquartile range 236–272) in the cat and 155 cells (range 104–203) in the monkey, compared with control levels of 35 cells (21–50) and 26 cells (18–33), respectively. Activation was predominantly in the ventrolateral area of the caudal PAG suggesting that the PAG is involved following trigeminally‐evoked craniovascular pain.

Activation of the Trigeminovascular System by Mechanical Distension of the Superior Sagittal Sinus in The Cat

Distension of dural sinuses in man produces migraine-like pain. In eight a-chloralose anaesthetized cats mechanical distension of the superior sagittal sinus with a small intraluminal device was used to activate single units in the dorsolateral C2 spinal cord. Units in this region have been shown to respond to electrical stimulation of the superior sagittal sinus in the cat model. Linked responses to mechanical dilatation could only be obtained with very rapid stretching stimuli or high amplitudes of distension of the vessel. Lower thresholds for transduction of distension in the vessel wall may depend on transferral to the dura or biochemical or neural pre-sensitization of the superior sagittal sinus. These data are consistent with the view that migraine is not primarily a vascular disorder but requires at least humoral or neural facilitation.