A prospective study on the role of the cerebellar (para)flocculus in tinnitus: the relation with electrophysiological parameters and inflammatory mediators

Tinnitus is the perception of sound without an external source. Chronic
tinnitus is prevalent among 5-15% of the general population, in 20% of cases
negatively affecting quality of life. No good treatment strategies are
available to this day, leading to a substantial economic burden to society. The
pathophysiology of tinnitus remains obscure. The cerebellar flocculus recently
received attention with regard to the pathophysiology of tinnitus in rats. In
rats, once tinnitus is established, the paraflocculus presumably becomes an
obligatory component in maintaining the condition.
In humans, the flocculus is located in the cerebellopontine angle (CPA).
Chronic tinnitus is experienced by 61% of patients after CPA tumor removal. The
flocculus is unavoidably manipulated during retrosigmoid removal of a CPA
tumor. Manipulation of the flocculus or compression by the tumor might
compromise the functions of the flocculus, leading to tinnitus. A retrospective
study in humans after surgical CPA tumor removal showed a positive correlation
between flocculus volume and tinnitus severity. Because the relation was
present on both the ipsilateral and contralateral side, it suggests a relation
between flocculus volume and tinnitus severity in general and therefore that
the flocculus plays a role in the regulation of tinnitus.
It has been noted during cerebellopontine angle (CPA) surgery that the
flocculus is sometimes firmly attached to the vestibulocochlear nerve. This
might be due to inflammation, as a common pathophysiological mechanism in which
two separate anatomical structures can glue together. Mean
neutrophil-to-lymphocyte ratio was higher in patients with severe tinnitus than
in controls and a correlation was present between serum levels TNF-α and
tinnitus loudness, suggesting the presence of inflammation as well. By
analyzing microglia activation and astrocyte presence in the choroid plexus and
cytokine analysis, it could be possible to objectify inflammation in the CPA
and particularly of the flocculus, which we hypothesize to lead to altered
function and tinnitus.
To conclude, this project investigates several aspects of flocculus function,
by determining tinnitus characteristics, by electrophysiological measurements
with an electrode placed directly on the flocculus during surgery, and by
exploring the possibility of inflammation.

Module 1
Primary Objective:
To objectify the relation between tinnitus and flocculus activity, quantified
by direct response measurements on the flocculus. Tinnitus is quantified by the
Tinnitus Functional Index score (TFI), tinnitus pitch and tinnitus loudness.

Secondary objective(s):
1. Is there a relation between floccular activity and flocculus manipulation?

Module 2
Primary Objective:
To examine the relation between tinnitus and inflammation, quantified by
expression of anti-glial fibrillary acidic protein (GFAP) as a marker of
astrocytes, anti-CD11b (OX-42) as a marker for microglia like cells and
cytokine levels in a choroid plexus biopsy, cytokine levels in serum and liquor
and serum inflammation levels.

This research protocol describes an observational study that will take place at
the University Medical Center in Groningen at the department of Neurosurgery.
Inclusion of patients is planned from February 17th, 2020 to September 30st,
2022, or until the target number of inclusions is acquired. Total follow-up is
6 weeks. Patients undergoing surgery for hemifacial spasms (microvascular
decompression) or a CPA tumor (retrosigmoid removal) are eligible for this
study. Patients who fulfill the inclusion and exclusion criteria and signed the
informed consent will be included. This study consists of two modules. All
patients will be asked to fill in a questionnaire regarding tinnitus and to
undergo audiometry and vestibular tests the day before surgery. Six weeks after
surgery patients will fill in the questionnaire again, which they receive at
home. They can sent the questionnaire to the UMCG using an UMCG envelope.

Patients can choose in which module(s) they want to participate.
Both modules are pilot studies. In module 1, we intend to objectify the
relation between tinnitus and flocculus activity, quantified by measuring
direct response measurements on the flocculus during surgery. In module 2, we
intend to examine the relation between tinnitus and inflammation, quantified by
expression of anti-glial fibrillary acidic protein (GFAP) as a marker of
astrocytes and anti-CD11b (OX-42) as a marker for microglia like cells in a
choroid plexus biopsy, cytokine levels in serum and liquor and serum
inflammation levels. A total of 20 patients with tinnitus and 20 patients
without tinnitus is aimed to be included in module 1 and module 2.

There are no benefits for patients to participate in this study. Possible risks
are minimal.
Vestibular test are obligatory measures for this study. Patients can become
nauseous or dizzy, but there is no risk for their health.
In module 1 electrodes will be placed on the flocculus during surgery. Because
of the chosen surgeries to take part in this module of the study, no additional
manipulation or interventions are necessary to obtain access to the flocculus.
The measurements on the flocculus itself are additional to standard patient
care. The same measurements have been done before at the UMCG without
complications (Kleinhuis et al., 2019).
In module 2 on the day of admission blood will be drawn, which is standard
care. During surgery, after induction of anesthesia and before incision, one
tube of blood will be drawn, using the arterial line if present. During surgery
liquor will be aspirated, which is standard care. Liquor aspiration is not
performed for diagnostic or therapeutic purposes, but to optimize visualization
during surgery by promoting brain relaxation. No additional procedures are
needed to obtain liquor. Additional to standard care, a biopsy of the choroid
plexus is taken during surgery. Because of the chosen surgeries to take part in
this module of the study, no additional manipulation or interventions are
necessary to obtain access to the flocculus. Taking a biopsy of the choroid
plexus is a known and save diagnostic method used in clinical practice for
diagnosis of suspected choroid plexus pathology for instance. Choroid plexus
extirpation and coagulation also used to be the standard treatment for
hydrocephalus. There is minimal risk of bleeding. This bleeding would be in
plain sight so it can be easily coagulated. A biopsy of the choroid plexus will
not be of influence on the homeostasis of the cerebrospinal fluid, because only
approximately 1% of the choroid plexus will be biopsied.
Based on information above, we can conclude that this study is a low risk study
(according to the NFU risk classification).