Olfactory event-related potentials and trigeminal event-related potentials –  first experience with objective olfactometry in the Czech Republic

Overview

Introduction: The clinical importance of olfactory evoked potentials (OERPs) and trigeminal evoked potentials (TERPs) in the diagnosis of olfactory disorders has recently increased. The advantage of the objective method is that OERPs are less biased than routinely used psychophysical olfactory tests. The clinical olfactometer gives precisely defined olfactory stimuli needed to elicit OERPs and TERPs. The principle of the method is based on the presentation of an odorant by a special device in the patient‘s nasal cavity and the registration of the brain response by electroencephalography (EEG) to the olfactory and trigeminal stimuli. For OERPs and TERPs, we evaluate the latencies and amplitudes of individual peaks and the N1–P2 interval. The absence of OERPs is a strong predictor of the presence of olfactory dysfunction. Aim of this study: These data will be a good basis for further research projects in the field of olfactory disorders. Materials and methods: Between 3/2021 and 6/2024, 187 subjects (99 females and 88 males) were enrolled in a prospective study. A clinical olfactometer OL 024 Burghart and an 8-channel EEG system OL 026 Burghart were used for measurements. Results: We present sample OERPs and TERPs curves from single groups of enrolled subjects. The 1^st group of normosmic subjects – healthy participants, subjects with a deviation of the nasal septum. The 2^nd group consists of subjects with chronic rhinosinusitis with nasal polyposis. The 3^rd group includes subjects after undergoing COVID-19. The 4^th group of enrolees are subjects with neurodegenerative disease, Parkinson‘s disease, and multiple sclerosis. The 5^th group includes subjects with tumors of the paranasal sinuses, olfactory region, and pituitary gland. The 6^th group consists of subjects with olfactory disorders who were indicated for examination for medico-legal reasons. The 7^th group consists of subjects with post-traumatic loss of smell. Conclusion: Sample OERPs and TERPs curves are demonstrated. The data obtained may very well be applied in the future as an internal guide for our other ongoing olfaction research studies. The absence of olfactory evoked potentials is a robust predictor of the presence of olfactory dysfunction. Objective olfactometry appears to be a method with increasing potential, especially in persons who have difficulty with commonly available psychophysical testing of the sense of smell, in patients with neurodegenerative disease, and in the medicolegal field.

Keywords:

olfactory event-related potentials – trigeminal event-related potentials – objective olfactometry – olfactory examination – odo-rant

Introduction

Psychophysical testing of the sense of smell currently plays a major role in daily clinical practice, but objective methods are needed whenever the cooperation of subjects in psychophysical tests is problematic. This may be the case, for example, in children, in persons with cognitive disorders or in the context of medico-legal examinations [1–3]. Thus, we have observed over the last 10 years the increasing clinical importance of the assessment of olfactory evoked potentials (OERPs) and trigeminal evoked potentials (TERPs) in the diagnosis of olfactory disorders. The advantage of the objective method is that OERPs are less biased than psychophysical olfactory tests. With increasing frequency, this diagnostic technique is used in clinical research and clinical practice to assess olfaction in individuals with olfactory disorders [1–3]. The clinical olfactometer gives precisely defined olfactory stimuli needed to elicit OERPs and TERPs. These evoked potentials (OERPs and TERPs) are an electrophysiological technique to assess changes in olfactory function and to objectively assess the integrity of the olfactory pathway [1]. The principle of the method is based on the presentation of an odorant by means of a special device in the patient‘s nasal cavity and the registration of the brain response by means of electroencephalography (EEG) to olfactory and trigeminal stimuli.

In the olfactory pathway, the electrical synaptic activity of neurons induced by various odorants is detected. The main advantage of this method is the objectification of the response to the odorant and the direct assessment of the preserved function of the olfactory nerve. This method can also help to detect misdiagnosis of the patient [2–4].

The clinical olfactometer works on the principle of injecting an odorous substance into clean, odourless, unpolluted air, which is fed through a tube to the edge of the patient‘s nasal cavity (Fig. 1).

All inner parts of the device must be made of materials that prevent contamination by other odours. The clinical olfactometer should be placed in a quiet and well-ventilated room. Substances that selectively stimulate only one of these are used to measure olfactory and trigeminal evoked potentials. Therefore, 2-phenylethanol (rose odor) is used as an odorant to selectively stimulate the olfactory nerve and CO₂ to stimulate the trigeminal nerve. The odorant is diluted in distilled water to form a suspension, which is bubbled through the air to create the vapor phase of the odor. This provides adequate humidity to prevent drying of the nasal mucosa during the experiment, and also maintains a constant temperature and prevents an unpleasant and undesirable trigeminal reaction. In the interval between stimuli, a humidified, warm, odourless air stream is conducted into the nasal cavity. When choosing a solvent, its physico-chemical properties must be taken into account. Differences in pH values or direct interactions between the liquid and the odorant may significantly alter their perception and therefore the measured values may not be valid. The resulting odorant vapor phase is then humidified (≥80%), heated to near body temperature (36 °C), and administered at a constant flow rate (8 litres per minute). Dry, too cold or too warm air, or higher flow rates irritate the end fibers of the trigeminal system, resulting in a sum of signals from the trigeminal and olfactory fibers [2, 3]. This also occurs during physiological changes in airflow in the nasal cavity. Therefore, the subject must breathe only through the mouth throughout the experiment. The vapor phase odorant mixture is delivered only by the device. If the air is too dry or cold, the nasal mucosa is congested. The appropriate study protocol is then selected in the clinical olfactometer‘s computer program. There, the duration of each stimulus, the intervals between stimuli, and the scheme in which the odorant and CO₂ are presented can be set. The aim is to select an order in which there is minimal risk of habituation to the odorant due to the alternation of odorant and CO₂, minimal stimulus duration to ensure adequate results, and minimal risk of patient harm [2, 3, 5].

The brain response to the odorant stimulus itself is measured using conventional EEG [6, 7]. Evoked potentials of OERPs and TERPs consist of a negative N1 component followed by a positive P2 component. The evaluation of the N1–P2 interval is an important parameter [6, 7]. We evaluate the latencies and amplitudes of individual peaks and the N1–P2 interval. The absence of OERPs is a strong predictor of the presence of olfactory dysfunction [3, 5, 8].

Aim of this study

To report the first information about this unique method in the Czech Republic, and to demonstrate sample curves of OERPs and TERPs in individual diseases associated with olfactory dysfunction. These data will provide a good basis for further research projects in the field of olfactory disorders.

Materials and methods

Our prospective study was approved by the local ethics committee of the Military University Hospital Prague, reference number: 108/16-49/2021 (project NU 22-09-00493), reference number 108/16-24/2021 (project MO 1012).

In the period 3/2021–9/2023, 187 subjects were included in the study. All participants signed the informed consent. The group consisted of 99 women and 88 men. The average age of the group was 42 years (range 18–84 years). The results of healthy subjects were statistically processed and published [9].

For the measurements we used a clinical olfactometer OL 024 Burghart, Germany (Fig. 1). This clinical olfactometer gives precisely defined odor stimuli that are necessary to elicit OERPs and TERPs. Subjects were stimulated with 2-phenylethanol (nervus olfactorius) and CO₂ (nervus trigeminus) according to standard procedures [1, 3]. Odorants were instilled into the left nostril as the standard. The resulting vapor phase of the odor was properly humidified (≥80%), heated to near body temperature (36 °C), and administered at a constant flow rate (8 l/min). During the experiment, white noise was generated in the subjects‘ ears using headphones at a volume of 60 dB. Patients were seated in a comfortable, relaxed position. They were asked to sit still, not to blink or swallow, and to breathe through their mouths. The test was performed in a well-ventilated room [1, 3, 5]. An 8-channel EEG system (OL 026; Burghart, Holm, Germany) was used to record responses.

Methods: OERPs were recorded at the top of the head (EEG, electrode Pz). 2-phenylethanol (50% v/v) was used to selectively activate olfactory afferents [3, 7, 9]. During the experiment, olfactory and trigeminal stimuli were presented separately. Each stimulus type was repeated 20-times and lasted 250 ms. The interstimulus time interval between each stimulus was 10–20 seconds.

TERPs: During the study, TERPs were recorded at the top of the head (EEG, electrode Cz). CO₂ gas (50% v/v) was used to selectively activate trigeminal afferents. During the experiment, olfactory and trigeminal stimuli were presented separately. Each stimulus type was repeated 20-times and lasted 250 ms. The interstimulus time interval between each stimulus was 10–20 seconds.

At the same time, the subjects underwent a psychophysical test of odorant identification (Sniffin‘ Sticks test) with a possible maximum score of 16. After presenting individual markers containing odorants, the subject was asked to correctly identify the substance from the four choices offered.

Subjects aged 18 years and older were included:

1. normosmia, healthy participants, and subjects with deviation of the nasal septum;

2. subjects with chronic rhinosinusitis with nasal polyposis with tissue sampling for the microbiome;

3. subjects after undergoing COVID-19;

4. subjects with neurodegenerative diseases, with Parkinson‘s disease in personal or family history, and subjects with multiple sclerosis;

5. subjects with tumors of the paranasal sinuses, olfactory region, and pituitary gland;

6. subjects with olfactory disorders indicated for examination for medicolegal reasons;

7. subjects with post-traumatic loss of smell.

The exclusion criterion was age below 18 years.

Results

We present sample OERPs and TERPs curves for the enrolled subjects in the Czech Republic. Here we present sample OERPs and TERPs curves in each group.

For simplicity and better understanding, we divide OERPs/TERPs into two basic groups:

• with present peaks N1 P2 as “present potentials”;
• with absent peaks N1 P2 as “absent potentials”.

Normative N1 and P2 wave data have been recently published in another one of our manuscripts and we refer to the part of the discussion where they are presented [9].

Group 1

Enrolled healthy normosmic subjects (N = 21); the normosmic subject with a nasal septal deviation, Sniffin‘ Sticks test (Identification section) 15 points, and sample curves with present potentials of OERPs and TERPs (Fig. 2a, b).

Group 2

Subjects with chronic rhinosinusitis with nasal polyposis (N = 50); the subject with chronic rhinosinusitis with nasal polyposis with tissue sampling for the microbiome, and sample curve with present potentials – OERPs/TERPs, in contrast with anosmia according to the Sniffin Stick identification test (identification of 1 point) (Fig. 3a, b).

Group 3

Post-COVID subjects (N = 71); the normosmic subject after COVID-19 associated with severe Guillain-Barré syndrome with present potentials of OERPs/TERPs (Fig. 4a, b), and Sniffin stick test – identification of 13 points.

In contrast, the subject after COVID-19 with a mild course and findings of absent potentials of OERPs/TERPs. The subject had six months of hyposmia and parosmia, and Sniffin stick identification test of 10 points (corresponding to hyposmia) (Fig. 4c, d)

Group 4

Neurodegenerative diseases (N = 39); the subject with Parkinson‘s disease with absent potentials of OERPs/TERPs, and Sniffin stick identification test of 0 points (anosmia) (Fig. 5a, b).

The subject with multiple sclerosis with present potentials of OERPs/TERPs, and Sniffin stick identification test of 14 points (normosmia) (Fig. 5c, d).

Group 5

Pituitary tumors (N = 3); subject after pituitary tumor surgery with sample curves of present potentials of OERPs/TERPs. Sniffin stick identification test of 12 points (normosmia) (Fig. 6a, b).

Group 6

Medico-legal examinations (N = 3); the first subject with post-COVID olfactory impairment acquired in the course of his occupation, referred for medico-legal reasons with present potentials of OERPs/TERPs, and Sniffin stick identification test of 4 points (anosmia) (Fig. 7a, b). The second subject with absent potentials of OERPs and with present potentials of TERPs, and Sniffin stick identification test of 4 points (anosmia) (Fig. 7c, d).

Group 7

Post-traumatic olfactory dysfunction (N = 3); the subject with post-traumatic olfactory loss with present potentials of OERPs/ TERPs. Sniffin stick identification test of 3 points (anosmia) (Fig. 8a, b).

Discussion

This new electrophysiological method of olfactory examination in the Czech Republic shows its use in practice for individual groups of diseases.

We have recently published pilot data on OERPs/TERPs in healthy normosmic subjects from the Czech Republic. We present our data in the table (Tab. 1) [9]. In a group of normosmic participants, we found statistically significant differences in amplitude by gender in TERPs. In OERPs, no statistically significant differences in amplitude, latency by age, or gender were found [9].

In subjects with nasal deviation, the curves of olfactory and trigeminal evoked potentials were the same as those of healthy subjects. In contrast, in chronic rhinosinusitis patients with nasal polyps, both absent potentials of OERPs/TERPs and present potentials of OERPs/TERPs curves were recorded. In this group, the importance of psychophysical testing of the sense of smell dominates. Evaluation of changes in OERPs/TERPs before/after administration of modern antibody therapy in subjects with chronic rhinosinusitis with nasal polyposis is expected in the near future.

The importance of evaluating OERPs and TERPs is unequivocal in neurodegenerative diseases such as Parkinson‘s disease, dementia, and multiple sclerosis [3, 10]. In subjects with Parkinson‘s disease, a frequent finding is a non-evaluable OERPs curve (absent potentials),

in contrast to the fact that present potentials of OERPs/TERPs are noted in subjects with a family history of Parkinson‘s disease.

Furthermore, the importance of OERPs and TERPs became more pronounced during the COVID-19 pandemic, when the number of patients with hyposmia and parosmia increased [7, 11]. The scientific outcome of the analysis of olfactory potential data after COVID-19 is expected, as only two publications have been published to date on the analysis of post-COVID curves of OERPs and TERPs. Chinese researchers have published data showing the importance of the duration of post-viral olfactory disturbance in relation to the return of olfaction after the disease. [5].

In subjects who had undergone COVID-19, we observed both variants with absent potentials of OERPs/TERPs and curves with normal values. Present potentials have also been described after undergoing severe post-COVID Guillain-Barré syndrome [7]. In contrast, there is an interesting example of absent potentials of OERPs/TERPs after COVID-19 in the subject who had documented OERPs/TERPs normal curves before undergoing COVID-19. The subject had hyposmia with parosmia six months after COVID-19 in which he perceived the smell of his favorite perfume as an unpleasant odor similar to the smell of human sweat.

In subjects after pituitary adenoma surgery, the OERPs/TERPs curves were present. This may be in agreement with the results in pituitary adenoma surgery published by the Czech authors, who reported that normosmia was present preoperatively in 93.7% of patients, and in patients with preoperative normosmia, postoperative hyposmia and anosmia were present in only 1.5% of the patients [12, 13].

Three subjects with olfactory impairment indicated for examination for medical-occupational reasons were also examined. All of them had anosmia after having undergone COVID-19 as part of their occupation as a surgeon and social worker. Sniffin stick psychophysical tests were suggestive of anosmia in the subjects. In contrast, OERPs/TERPs curves were present in two subjects and absent in one subject. This is a highly topical tricky issue that will be further developed in its own line of research.

According to Chinese authors, in the near future, electrophysiological olfactory assessment with evaluation of OERPs/TERPs should become the gold standard in olfactory investigation [5]. We, like the team of Dresden smell and taste researchers led by Professor Thomas Hummel think that this is an optimistic perspective [1, 9]. We believe that psychophysical olfactory tests will be the gold standard for a long time to come for several reasons: 1. the purchase price of a clinical olfactometer with 8-channel EEG is relatively high; 2. only a few departments in the world have the equipment; 3. the method requires working with a med-tech engineer and is equally demanding to service [1, 9, 14–16]; 4. psychophysical methods allow examination with various odorants and it is the high degree of qualitative assessment of odorants that is very important for olfaction; 5. psychophysical methods allow examination not only of identification but also of threshold, discrimination, and liking of odorants; and 6. psychophysical methods require less time.

Moreover, the aim of using OERPs/ /TERPs is not to supplement the psychophysical examination of the sense of smell, but to add subjective olfactory methods and to apply them in practice, especially in medico-legal matters or in situations where the patient is unable to undergo an olfactory examination. The case reports of olfactory disorders following COVID-19 then provide examples where patients suffer from anosmia on the basis of odorant identification testing, but OERPs are present. Therefore, Professor Hummel‘s team introduced the term functional anosmia to reflect significant olfactory impairment (anosmia upon psychophysical examination). In some patients with functional anosmia, some olfactory stimuli may be preserved (OERPs are present), but the sense of smell is not useful in practical life [17]. Moreover, OERPs do not provide a more accurate picture of central processing of the olfactory stimulus. In psychophysical examinations, the peripheral function of the olfactory organ corresponds very well to the threshold examination, whereas the central processing of the olfactory stimulus is important in the evaluation of discrimination, identification, or liking of odorants. In studies, another objective method, functional magnetic resonance imaging, is then used to more accurately describe the central processing of odor stimuli.

Conclusion

Our analyzed OERPs and TERPs pilot curves will be used as an internal guide for our other ongoing olfaction research studies. The assessment of the presence of OERPs will be an important parameter for the evaluation of olfactory disorders. The absence of olfactory event-related potentials is a strong predictor of the presence of olfactory dysfunction. Detailed examination of the sense of smell should not be neglected.

Electrophysiological olfactory assessment appears to be a method with potential for future research on olfaction, especially in persons who find it difficult to manage commonly available psychophysical testing of the sense of smell, in patients with neurodegenerative diseases, and in the medico-legal field. Further research into objective olfactory measurement and data collection in practice should lead to the widespread use of objective olfactometry across medical disciplines in the future. Psychophysical examination of the sense of smell, OERPs, and TERPs are methods that are highly complementary and provide us with a very detailed picture of the function of the olfactory analyzer.

Funding

This study was funded by Project NU 22-09- -00493 and Project NW 24-08-00324 of the Ministry of Health of the Czech Republic and by Project MO 1012 of the Ministry of Defence of the Czech Republic.

Conflict of interest statement

The author of the article declares that he has no conflict of interests in connection with the topic, creation and publication of this article, and that neither the creation nor the publication of the article was supported by any pharmaceutical company. This statement also applies to all co-authors.