Data from sixty-two randomly chosen healthy young males, aged 18–31 years (mean 24.87 y, SD ± 3.74 y), were taken into account in the study. Thirty-four of them were smokers and 28 non-smokers. None of them had received medication for any disease during the previous six months. No subject suffered from otolaryngological diseases, acute or chronic, such as rhinitis and sinusitis. Oral hygiene was also examined before testing. Concerning smokers, the duration of smoking was between 1 and 6 (3.2 ± 0.7) years and the number of cigarettes smoked ranged between 12 and 40 (18 ± 2.3) per day. All the smokers held their cigarettes, when smoking, at the centre of their lips. All the subjects were soldiers serving at the same military unit during the last three months and had similar nutritional habits. The majority of the examined persons were right-handed and only few of them (3 non-smokers and 1 smoker) were left-handed. They participated in the study only after they had been informed of its background and purpose and after their written consent was obtained. The protocol was reviewed and approved by the Institutional Review Board of The Aristotle University of Thessaloniki. Measurements were conducted according to the Helsinki Declaration.
In order to minimize variations in technique and interpretation of results, all examinations were carried out by the same researcher (PP). Before being tested, all the subjects were asked whether if they were experiencing any abnormal taste at the time. The checklist for recording their history is shown in Table .
Checklist for history taking in patients complaining of taste problem
Taste sensitivity was evaluated with EGM. Electrical stimuli were delivered with an electrogustometer (TR-06, Rion Co, Japan) with a single, flat, circular stainless steel stimulus probe (5 mm diameter). The apparatus produces stimuli of low current and known durations (0.5, 1, 1.5 and 2 seconds). A feedback circuit controls the output current with an error of < 1% [18
]. The thresholds varied with increases in the duration, though in many subjects the measurements concerning 1 and 1.5 seconds were the same.
All subjects had been instructed not to drink anything for an hour before the start of testing. Before measuring thresholds, a stimulus of 30 dB was administered to ensure that every subject could recognize electrogustometric stimuli. Stimulation started at the lowest stimulus strength (-6 dB) and increasingly stronger stimuli were presented until the subject recognized the stimulus. If the threshold for stimulus perception was not clearly distinguished, the next higher- and lower-strength stimuli were presented.
The electric threshold scores were measured at points on the right and left sides of the tongue apex (2 cm from the apex), the vallate papillae and the soft palate. Among healthy population electric thresholds for the apex, vallate papillae and soft palate are considered to be up to 8, 14 and 22 dB respectively [18
]. Stimuli of all the available durations (0.5, 1, 1.5 and 2 seconds) were applied in order to investigate whether the different duration affected the recorded thresholds. Electric stimuli were applied, beginning at -8 dB to 34 dB (3–400 μÁ) in 2 dB steps. The relationship between the logarithmic control settings and the output currents is shown on Tables and .
Output Current dB Readings and Output Current
Output Current dB Readings and Output Current
We started measuring taste thresholds from right to left side. The threshold of the right side of the soft palate is Threshold A
, the one of the vallate papillae is Threshold B
and the right side of the tongue apex Threshold C
. The corresponding thresholds of the left side of the tongue are Threshold D
, Threshold E
and Threshold F
(F for the left side of the soft palate). Cases where the threshold could not be measured were assigned as threshold value of 36 dB [19
]. All 6 sites were tested with the same stimulus duration before proceeding to the next duration. In that way an interval of 3–4 minutes took place before the stimulus of the next duration was applied on a site, leading to a lower possibility of adaptation. The subjects had been instructed to recognize whether they perceived a sour/metallic taste suggesting gustatory function (taste threshold) or an electrical sensation suggesting trigeminal function.
Imaging was performed using a 30° contact endoscope (CE; magnification, × 60 and × 150; Karl Storz, Tuttlingen, Germany). Identification of fPap was performed at first by a noncontact technique. Subjects had rinsed their mouth with water prior to imaging. A contact technique was used without staining for imaging of subepithelial vessels. Methylene-blue 1% was used afterwards to stain epithelia and taste pores. Application of methylene blue was preceded by careful suctioning of the mucus. A filter paper strip delineating an area of 1 cm2
was placed in a paramedian position on the tongue tip [20
The subjects opened their mouth and the CE was placed on the methylene-blue stained surface of the tongue. To resolve the instability of the tongue during imaging the subjects were advised to hold the tip of the tongue with the upper and lower teeth, not too strongly in order to avoid stasis and hypaeremia which could lead to wrong estimation. They were also seated in a chair with their head and neck supported by a neck pillow. The form of the papillae and blood vessels were classified according to a previous classification [21
]. The forms of the papillae were classified in Type 1
, (egg-shaped or long ellipse type -No surface thickness), Type 2
(slight thick surface as compared to type 1), Type 3
(thick and irregular surface) and Type 4
(remarkably flat and atrophic surface). The classification of blood vessels was Type A
(clear loop and wooden branch shape), Type B
(unclear loop and wooden branch shape), Type C
(elongated blood vessels), Type D
(granular shape or dotted shape) and Type E
(unclear blood vessels).
For estimating the density of fPap the best image from every individual was used. fPap could be distinguished from filiform papillae (which were stained dark), by their very light staining [22
The results were analysed with SPSS 12 for Windows (SPSS Inc. Chicago, IL, USA).
The null hypothesis was that there was not a statistical difference between the subjects of the two groups. Non-parametric tests were applied because there was no normal distribution of the assessed thresholds. The level of statistical significance was set to p < 0.05. The thresholds of the two groups were compared using Kruskal-Wallis and Mann-Whitney tests. The Bonferroni correction was used where it was necessary. The same tests were used for the comparison between the thresholds recorded in each stimulus-duration in both groups.